RNA interference (RNAi) screens have enabled the systematic analysis of many biological processes in cultured cells and whole organisms. The success of such screens and the interpretation of the data depend on the stringent design of RNAi libraries. We describe and validate NEXT-RNAi, a software for the automated design and evaluation of RNAi sequences on a genome-wide scale. NEXT-RNAi is implemented as open-source software and is accessible at http://www.nextrnai.org/.
RNA interference (RNAi) screens have become an important tool for the identification and characterization of gene function on a large-scale and complement classic mutagenesis screens by providing a means to target almost every transcript in a sequenced and annotated genome. RNAi is a post-transcriptional gene silencing mechanism conserved from plants to humans and relies on the delivery of exogenous short double-stranded RNAs (dsRNAs) that trigger the degradation of homologous mRNAs in cells [1, 2]. As an experimental tool, RNAi is now widely used to silence the expression of genes in a broad spectrum of organisms .
The availability of genome-wide RNAi libraries for cell-based assays and whole organisms has opened new avenues to query genomes for a broad spectrum of loss-of-function phenotypes [4, 5]. The number of sequenced genomes is steadily rising, enabling reverse genetic approaches using RNAi in many novel model systems, including, for example, the medically relevant vector Anopheles gambiae and species used to study evolutionary aspects of development, such as Tribolium castaneum, Acyrthosiphon pisum and Schmidtea mediterranea. RNAi libraries will facilitate the functional characterization of genes in these species, either through studying smaller subsets of candidates or on a genomic scale.
The design of RNAi reagents is key to obtaining reliable phenotypic data in large-scale RNAi experiments. Several recent studies demonstrated that the degradation of non-intended transcripts (so-called 'off-target effects') and knock-down efficiency depend on the sequence of the RNAi reagent and have to be carefully monitored [6–13]. Based on experimental studies, rules for the design of RNAi reagents have been devised to improve knock-down efficiency and simultaneously minimize unspecific effects.
In invertebrates such as Caenorhabditis elegans and Drosophila, RNAi can be triggered by long dsRNAs that are intracellularly broken down into short interfering RNAs (siRNAs) [1, 14, 15]. The design of a long dsRNA therefore needs to take into account both the properties of the target sequence, for example, its sequence complexity, as well as the properties of all siRNAs contained within the long dsRNA, such as their predicted target specificity and efficiency. Because long dsRNAs are often generated by in vitro transcription, the design of suitable primer pairs to amplify in vitro transcription templates through PCR from genomic DNA or cDNAs must be implemented.
In contrast, RNAi-mediated silencing in mammalian cells is achieved through siRNAs of 21 to 23 nucleotides  to circumvent the activation of an interferon response . Such short dsRNAs can be generated by different methods. For mammalian cells, vectors transcribing short-hairpin RNAs [18–20] or synthetic siRNAs  are commonly used. Several recent studies have highlighted favorable sequence characteristics and suitable chemical modifications for these reagents [21–23]. The design of long in vitro endoribonuclease-prepared siRNA reagents (esiRNAs)  resembles that of long dsRNAs for model organisms. We list several factors that are important for the design of RNAi reagents in Figure 1.
Quality control parameters for RNAi reagents at different stages of the design pipeline. (a) Long dsRNAs that have regions of low complexity, for example, CA[ATCG] repeats or simple nucleotide repeats, can exert unspecific and cytotoxic effects. The quality of the primer designs used to synthesize amplicons from DNA sources is crucial, in particular when the synthesis is performed in 96- or 384-well formats where primers should have similar melting temperatures. (b) Dicer-mediated cleavage of long dsRNAs leads to the generation of siRNAs of lengths between 19 and 23 nucleotides . The quality of siRNAs depends on their ability to efficiently enter the RNA-induced silencing complex (RISC) and to access the target mRNA. This is influenced by thermodynamic properties, base preferences and chemical modifications. The specificity of siRNAs is influenced by sequence-independent and sequence-dependent features. siRNAs can trigger interferon responses or show concentration-dependent cytotoxic effects, independent of their sequence. Silencing of unintended target transcripts can occur through perfect and imperfect sequence homologies to the siRNA and through 'seed matches' to the transcript 3' UTRs. See text for details.
For large-scale functional screens, the design of RNAi reagents is particularly important because the specificity and efficiency of individual RNAi reagents can rarely be validated on a genome-wide scale. The systematic application of design criteria, often on poorly defined gene models, has a direct impact on the expected false positive and false negative rates of phenotypic screens. Previous computational tools are available to design short and long dsRNAs for individual genes [12, 25, 26]. However, the systematic and reproducible design of RNAi reagents for large sets of genes or even whole genomes using an expanded set of parameters, such as target analysis for all splice isoforms, overlap analysis with SNPs and calculation of seed match frequency, has remained an unresolved issue.
Here we present NEXT-RNAi, a software tool for the design and evaluation of RNAi libraries that can be used for projects with targets ranging from a limited gene set to a whole-genome scale. NEXT-RNAi can process annotations from various sources and thereby provides a powerful RNAi design pipeline for virtually any genome that is available in public databases. NEXT-RNAi can also be used to design independent RNAi reagents to complement existing libraries. To demonstrate its flexibility, we have designed multiple genome-wide RNAi libraries for different organisms, including Drosophila, Anopheles, Tribolium and humans. NEXT-RNAi also offers the opportunity to automatically evaluate and re-annotate existing RNAi libraries by generating user-friendly reports to reflect the regular update of genome annotations.
To validate knock-down efficiency of NEXT-RNAi's reagent designs, we generated two independent sets of long dsRNAs targeting protein and lipid phosphatases expressed in Drosophila D.Mel-2 cells and verified transcript knock-down by quantitative real-time RT-PCR.
RNAi screens rely on the design of large-scale libraries comprehensively covering annotated transcriptomes. The design of RNAi libraries requires the identification of suitable target regions that minimize the potential for off-target effects, increase the silencing capacity and allow an efficient synthesis of the reagents. Often, multiple independent designs that meet these requirements are used to confirm RNAi-induced phenotypes.
Figure 2 illustrates the workflow of NEXT-RNAi for the automated design and evaluation of RNAi reagents (see also Additional file 1), and Figure 3 exemplifies the steps typically performed for the design of a long dsRNA. The input target sequences (Figure 3a) are first analyzed for regions of low complexity that have been shown to exert promiscuous off-target effects . NEXT-RNAi identifies tandem trinucleotide repeats of the type CA[ACGT] (CAN) and can also use the mdust  filter program (with default parameters) to find, for example, simple nucleotide repeats or poly-triplet sequences other than CAN (Figure 3b). The function of the intracellular Dicer protein  is then simulated by computationally 'dicing' the input target sequences into all possible siRNAs with a (default) length of 19 nucleotides. siRNAs may cause unspecific gene silencing via short stretches of homology with unintended mRNAs [27, 30, 31] or by a route similar to miRNA-mediated silencing through sequence similarity in positions 2 to 7 or 2 to 8 of the siRNA guide strand to the 3' UTR of unintended transcripts [32, 33]. NEXT-RNAi assesses the specificity of siRNAs by mapping them to the transcriptome. An siRNA is considered 'specific' if only isoforms of the same gene are targeted (with perfect homology; Figure 3b). The number of siRNA seed matches (seed complement frequency) is determined by mapping all the unique seeds to a user-defined database containing, for example, 3' UTR sequences. Several criteria can be taken into account to determine the predicted efficiency of an siRNA, including asymmetric thermodynamic properties [8, 10], G/C content, structural properties  and base preferences at several positions [6, 9, 11]. NEXT-RNAi implements two scoring methods to assess the predicted siRNA efficiency, here referred to as the 'rational'  and 'weighted'  methods. Scores range between 0 and 100 (Figure 3b). A previous analysis by Reynolds et al.  reported that siRNAs with efficiency scores ≥66.7 (on our normalized scale) were efficient silencers in human cells; and Shah et al.  found that designs with scores ≥63 were efficient. Analysis of 2,431 knock-down validated siRNAs (from Huesken et al. ) for their predicted efficiency (Additional file 2) shows a good correlation between the normalized inhibitory activity of the siRNAs and the predicted efficiency score (correlation of 0.52 and 0.51 for the 'rational' and 'weighted' methods respectively; P-value <2.2e-16).
Overview of the NEXT-RNAi workflow. NEXT-RNAi requires a defined set of input files in FASTA or tab-delimited formats. First, the program filters the input target sequences for six (default) or more contiguous CAN repeats and for other regions of low complexity (for example, simple nucleotide repeats) using mdust. Sequences are then 'diced' to generate all possible siRNA sequences with a default length of 19 nucleotides (nt) and an offset of 1 nucleotide. Subsequently, each siRNA is mapped to a user-defined off-target database (for example, the whole transcriptome) with Bowtie  to determine its specificity. The specificity is set to one if the siRNA targets a single gene or to zero otherwise. In the next step, the predicted efficiency of each 19-nucleotide siRNA is computed. Two methods can be selected, the 'rational' method according to Reynolds et al.  and the 'weighted' method according to Shah et al. , assigning each siRNA an efficiency score between 0 and 100. Optionally, the seed complement frequency for each siRNA can be computed for any FASTA file provided (for example, a file containing 3' UTR sequences). siRNAs that did not pass the low-complexity filters, show perfect homology to multiple target genes or do not meet the user-defined cutoffs for efficiency or seed complement frequency are excluded from the queried target sequences. Remaining sequences are used as templates for primer design (with Primer3 ) for long dsRNAs or are directly subjected to the final ranking for the design of siRNAs. Designs are ranked by (i) their predicted specificity and (ii) their predicted efficiency and, in the case of siRNA designs, (iii) their calculated seed complement frequency. Sequences can also be evaluated for additional features, such as homology to unintended transcripts, or SNP and UTR contents. Final designs can be visualized using GBrowse . All results are presented in a comprehensive HTML report and are also exported to text files.
Example of design and filter methods applied by NEXT-RNAi. (a) Visualization of the Paf-AHalpha gene model and transcripts. Regions labeled as 'common region' serve as input for NEXT-RNAi (ORF = open reading frame, UTR = untranslated region). (b) Quality measures computed by NEXT-RNAi for the common regions. Blue and red regions label predicted low-complexity regions (including CAN repeats) and 19-nucleotide off-target regions, respectively. The lower panel shows the predicted siRNA efficiency according to Shah et al.  (averaged for ten siRNAs). (c) NEXT-RNAi predictions of optimal target sites (green) after discarding 19-nucleotide off-target and low complexity regions and regions <150 nucleotides or >250 nucleotides. If available, these regions are directly used as templates for primer designs (left and middle panels). Otherwise, a redesign method is used that connects closest 'optimal' neighbors until a region suitable for primer designs is identified (right panel). Potential dsRNAs are finally ranked by their specificity and efficiency.
All quality parameters measured prior to this step, including the prediction of specificity, efficiency and low complexity, are applied as filters on the input sequences to identify optimal RNAi target sites. The set of filters can be further expanded by also including cut-offs on the seed complement frequency and sequence filters on conserved miRNAs seeds (for each siRNA). For the design of long dsRNAs, Primer3  is then used with user-defined settings to design primer pairs required for the PCR during dsRNA synthesis (Figure 3c). In case the optimal target sites identified are too short for designing primers (colored grey in Figure 3c), NEXT-RNAi implements a redesign routine that can be enabled by the user. This routine identifies those optimal target sites that are closest to each other and combines them by including the 'suboptimal' region in between. This step is carried out iteratively until the region is long enough for designing primers (for example, see right panel in Figure 3c).
Long dsRNA or siRNA designs are finally ranked by predicted specificity and predicted efficiency. For the ranking of siRNAs, designs with low seed complement frequency are prioritized. Since long dsRNAs contain many different siRNAs, two efficiency scores are reported: the average efficiency score of all contained siRNAs and the absolute number of efficient siRNAs (efficiency above a user-defined cutoff). The user-defined number of top-ranked designs for each target can be evaluated further by mapping them to the genome (with Bowtie  or Blat ), by determining the overall homology to other transcripts (with Blast ) or by calculating the overlap with other sequence features such as SNPs or UTRs.
NEXT-RNAi outputs design information in a tab-delimited text file and generates a comprehensive HTML report including a graphical display of designs in GBrowse  (Additional files 3 and 4). Further, details are available in FASTA, GFF (generic feature file) and AFF (annotation file format) formats for additional sequence analyses and straightforward reagent visualization in any genome browser.
We next set out to apply the software to design novel genome-wide RNAi libraries for different organisms, including Drosophila melanogaster, T. castaneum, A. gambiae and Homo sapiens.
Several RNAi libraries for cell-based  and in vivo RNAi screens (Vienna Drosophila RNAi Center (VDRC) , Fly stocks of National Institute of Genetics (NIG-Fly) and Transgenic RNAi Project (TRiP) libraries) have been constructed covering almost all genes annotated in the Drosophila genome. We used NEXT-RNAi to design multiple independent long dsRNAs targeting all Drosophila genes based on the latest genome release (FlyBase  release 5.24) in one run. Each design targeted all splice variants of a given target gene. To this end, we computed regions common to all annotated isoforms of the 14,898 coding or non-coding Drosophila genes. To further increase the number of potential target sites, we split common regions longer than 700 nucleotides into two sequences of equal length. This resulted in 74,907 common regions overall or an average of five regions per gene to be used as input for NEXT-RNAi. The Drosophila transcriptome was used as a database to evaluate the siRNA specificities ('off-target' database). NEXT-RNAi design options were adjusted to exclude low-complexity regions, CAN repeats, 19-nucleotide siRNA matches to unintended transcripts and siRNAs containing miRNA seeds (as predicted by miRBase ). The length-window for long dsRNA designs was set to 80 to 250 nucleotides. We included an iterative redesign, as described above, for sequences initially failing to meet these criteria. The best design for each input sequence was further evaluated for homologies (Blast E-value <1e-10) to unintended transcripts and for overlaps with UTRs.
The NEXT-RNAi output is exemplified in Additional files 3 and 4. Summarized results for the designs are presented in Additional file 5. The full report is available on our companion website . In total, 70,149 designs were calculated, covering 99.4% of all annotated genes with at least one dsRNA and 88.7% with multiple independent designs. Eighty-three gene models could not be targeted because of gene-spanning low complexity regions. Each gene model was, on average, targeted by 4.7 independent designs, 90.7% of which lack any perfect homology to any location other than the intended target transcripts of more than 18 nucleotides. In some cases, dsRNAs including 19-nucleotide matches could not be avoided, for example, for paralogous gene families with high sequence similarities or long overlaps (for example, actin or histone families).
A similar approach was used to generate independent designs for all predicted exons included in the 'official gene set' (available from BeetleBase ) of the recently sequenced genome of the red flour beetle, T. castaneum . Tribolium has become an important model organism for developmental and evolutionary studies, and efficient RNAi through injection of long dsRNAs has been demonstrated .The newly designed RNAi reagents covered 99.4% of all predicted gene models (83.2% with multiple independent designs), of which 92.9% lacked any predicted 19-nucleotide off-targets (Additional file 5).
The mosquito A. gambiae is widely studied to analyze the mechanism of innate immunity as a vector for Plasmodium falciparum. RNAi by long dsRNAs has been demonstrated in vitro and in vivo and leads to efficient depletion of mRNAs . Based on VectorBase  annotations, we designed RNAi reagents covering 95% of all genes (90.1% of all genes were covered by independent designs). Of all the designs, 89.2% had no unintended 19-nucleotide match in the Anopheles transcriptome (Additional file 5).
RNAi experiments in mammalian systems require the application of either in vitro-diced long dsRNAs (esiRNAs) or synthetic siRNAs. Here we designed reagents for both approaches to target all human genes annotated by the National Center for Biotechnology Information (NCBI) RefSeq database  (Additional file 5). Regions common to all RefSeq transcripts of the same gene were computed for all human genes and used as target sites for multiple independent esiRNA and siRNA designs per gene. Although both libraries covered almost the entire genome (esiRNAs, 97.8%; siRNAs, 99.9%), siRNA designs allowed a higher coverage and targeted more genes without predicted 19-nucleotide homologies to unintended transcripts (83.4% (siRNA) compared to 73.8% (esiRNAs) of the genome). The mean of predicted efficiency scores for siRNA designs was 84.76 (the 'weighted' method was used with a cutoff of 63), about 39% of the designs have low seed complement frequencies (less than 1,000 seed matches; RefSeq annotated 3' UTRs were used for seed match computation) and about 12% of the siRNAs contain annotated SNPs (from dbSNP ), which can interfere with siRNA function.
The complete description and NEXT-RNAi reports of the libraries for different organisms are available at . Similarly, NEXT-RNAi could be applied to other recently sequenced genomes, including Schmidtea mediterannea and Acyrthosiphon pisum, for which RNAi has become the method of choice for functional experiments.
A challenge for the interpretation of screening experiments is the correct annotation of available RNAi reagents; this includes the assessment of quality control parameters, their mapping to the genome and updating their target information for new genome annotation releases.
NEXT-RNAi enables the re-calculation of specificity, efficiency and other features of libraries of long dsRNAs and siRNAs. As examples, we performed a re-annotation of eight large-scale RNAi libraries designed for the Drosophila genome (Ambion, Heidelberg 2 (HD2), Heidelberg Fly Array/Drosophila RNAi Screening Center (DRSC) v1.0 , DRSC v2.0 , OpenBiosystems v1/v2, Medical Research Council (MRC), NIG-Fly and VDRC ) using the FlyBase annotations of release 5.24 (Additional file 6). With the exception of the HD2 and DRSC v2.0 libraries, all libraries covered less than 90% of the genome. This might result in part from the fact that they were designed for previous genome releases (release 3 or earlier). A comparison between the libraries showed how the design strategies evolved over time. While the designs of the HD2 and DRSC v2.0 libraries avoided both 19-nucleotide off-target effects (26.6% and 31.1% of all dsRNAs in HD2 and DRSC v2.0, respectively) and CAN repeats (0.5% and 1.8% of all dsRNA in HD2 and DRSC v2.0, respectively), older libraries, including DRSC v1.0 and MRC, contain a significantly higher percentage of dsRNAs with predicted 19-nucleotide off-targets (37.1% and 51.5%, respectively) and CAN repeats (5.3% and 5.4%, respectively). NEXT-RNAi also allows for the assessment of further parameters of the reagents. In this analysis, we computed the number of siRNAs with known miRNA seeds (from miRBase ) contained within each long dsRNA. With an average of 1.9, the Ambion library contains the fewest miRNA seeds per dsRNA, potentially because Ambion dsRNAs are rather short (255 nucleotides). Analyzing long dsRNAs for overlaps with UTRs reveals that designs in the DRSC v1.0, MRC, NIG-Fly and VDRC libraries were aimed at targeting open reading frames only (in all of these libraries, less than 8% of the reagent targets predicted UTRs).
An important experimental step during the confirmation of candidate genes from RNAi screens is the validation of phenotypes with independent designs . We used NEXT-RNAi results to identify the number of genes that could be targeted with independent designs through pairwise combinations of all Drosophila RNAi libraries (Figure 4; Additional file 7; complete reports are available for download). Pairwise combinations of the Ambion, DRSC v2.0 and HD2 libraries provide the highest number of independent reagents (for example, 6,623 genes covered by HD2 are covered by at least one independent design in DRSC v2.0). Some libraries overlap to a large extent and would be less advisable to use for confirmation screening. For example, combining the DRSC v1.0 and MRC libraries covers only 2,593 genes by independent designs. The analysis done provides also a helpful resource to identify in vivo RNAi lines of VDRC and NIG-Fly libraries that can be used for confirmation experiments with a second, non-overlapping dsRNA.
Pairwise comparison of Drosophila RNAi libraries. Eight Drosophila RNAi libraries are compared to identify the number of genes that are targeted by multiple libraries (outer ring, light grey) and the number of genes that are targeted by independent designs (inner ring, dark grey). The reference for the ring sizes is the combination of libraries commonly targeting the most genes (HD2/DRSCv2: 13,197 genes).
We also re-annotated human siRNA libraries from Ambion (Silencer Select Library) and Qiagen (human druggable v3.0 and human whole genome supplement v1.0), containing 64,781 and 70,308 siRNAs, respectively (Additional file 6). Of all siRNAs in the Ambion and Qiagen libraries, 3.4% and 10.1%, respectively, lacked any annotated target gene in NCBI RefSeq release 40; 84.2% and 92.4% show perfect homology to a single target gene; and 5.7% and 4.1% perfectly match multiple targets. The libraries cover 75% and 65.3% of all currently annotated NCBI and Entrez genes, respectively. About 9% of siRNAs in both libraries contain annotated SNPs (from dbSNP). More than one-third of the siRNAs in the Qiagen library overlap with annotated UTRs in their target transcripts (by at least one base), but only about one-tenth of the siRNAs in the Ambion library do so. Libraries also differ in the mean of predicted siRNA efficiency scores (using the 'weighted' method), with 74.65 for the Ambion and 57.58 for the Qiagen library. Of the Ambion and Qiagen siRNAs, 9.8% and 5.3%, respectively, have low seed complement frequencies (less than 1,000 seed matches in RefSeq annotated 3' UTRs).
To validate the knock-down efficiency of reagents designed by NEXT-RNAi, we designed two independent long dsRNAs (see Additional file 8 and companion website for details on the design) for all Drosophila protein- and lipid-phosphatases expressed in D.Mel-2 cells (Gene Expression Omnibus (GEO) accession [GEO:GSE21283]). We found 49 phosphatases expressed at five or more RPKM (reads per kilobase gene per million reads; Additional file 9). The reagents were synthesized using a two-step PCR procedure followed by in vitro transcription  with a 100% synthesis success rate.
After RNAi knock-down for 5 days (Figure 5a), transcript levels were determined using quantitative RT-PCR. Out of 98 dsRNAs, 87 (88.8%) caused a decrease in mRNA levels of more than 60%; half of the dsRNAs achieved a knock-down exceeding 80% (Figure 5b; Additional file 10). Eleven mRNAs showed little or no knock-down, six of which could not be detected reproducibly in this assay. For 37 of the 49 genes, we found that both independent designs decreased mRNA levels by at least two-thirds. For eight genes, only one design and for four genes, no designs could be validated with this knock-down strategy (Figure 5c).
Knock-down validation of Drosophila phosphatases. (a) Experimental workflow for knock-down validation. (b) Frequencies of observed knock-down efficiencies for independent designs. (c) The number of genes efficiently silenced (knock-down >66%) by both independent designs, only one or neither design. qPCR, quantitative RT-PCR.
Overall, our results show that NEXT-RNAi designs efficiently silenced targeted mRNAs. Furthermore, the independent designs led to highly reproducible knock-downs (Pearson correlation coefficient of 0.85), indicating that the observed depletion efficiency depended on the targeted mRNA rather than differences in the NEXT-RNAi designs.
In large-scale RNAi experiments, the design of genome-wide silencing libraries has remained an important problem due to the flux of gene annotation and novel insights into the mechanisms that influence RNAi efficiency and off-target effects. We present an approach for the rapid design of whole-genome RNAi libraries and the re-annotation of already existing reagent collections. The method is flexible, identifies multiple independent reagents per gene model and has been implemented in an organism-independent manner. The design process is fully automated and can use annotations from various sequence- or model-organism databases as input, thereby enabling the design of RNAi reagents for any sequenced (and annotated) organism.
We have designed several independent RNAi libraries for a diverse group of organisms. The automated pipeline yielded designs for more than 95% of all predicted genes in the first round of prediction. All library designs are available as a resource for download from our webpage . We validated the knock-down of 98 long dsRNAs directed against 49 Drosophila phosphatases expressed in our tissue culture model and found that approximately 89% of the reagents caused at least 60% mRNA knock-down. The application of a standardized design pipeline for independent designs leads to reproducible knock-downs in our experiments (correlation of 0.85 between the independent designs).
RNAi screens have become a key tool for functional genomic analyses. The interpretation of the increasing number of published data sets obtained through RNAi screens relies heavily on correctly annotated reagents. Phenotypes derived from large-scale screens should be linked to the sequence of the RNAi reagent rather than the gene model because off-target or splice-variant-specific silencing can rarely be excluded. For the correct interpretation of RNAi screens, and also the comparison between different libraries, reagent-to-gene-model linkages must be re-mapped in regular intervals because most genome annotations are still in flux. NEXT-RNAi can be used to rapidly evaluate and re-annotate existing genome-wide libraries. For example, we have applied the algorithm to re-annotate RNAi libraries for Drosophila and human cells. Our analysis of eight genome-wide RNAi libraries for Drosophila revealed differences in genome coverage and predicted quality (for example, specificity), most likely depending on two factors: the quality of the underlying genome release and the factors known to influence reagent quality at the time of the library design. Further, reagents in these libraries often share target sites, thus preventing an independent confirmation of phenotypes on a genomic scale. The re-annotation of commercially available human libraries revealed that a substantial part of the siRNAs (Ambion library, 15.8%; Qiagen library, 7.5%) either do not target the intended gene or are predicted to silence additional loci, demonstrating that quality control at the level of sequence mapping is crucial for the interpretation of large-scale screens.
Several tools for the design of RNAi reagents exist (including, for example, E-RNAi , DEQOR , SnapDragon , and siR, and commercial design tools such as siDESIGN Center (Dharmacon, ThermoScientific), BioPredsi (Qiagen) and siRNA Target Finder (Ambion)). However, these tools can only be used for designing long dsRNAs or siRNAs on a gene-by-gene basis. In contrast to available tools, our method allows for rapid batch design and evaluation of RNAi libraries for complete genomes or for any defined set of genes. In addition, our approach uses multiple parameters to calculate or evaluate designs, including sequence complexity, efficiency and specificity indicators, and allows for further refinement by scoring overlap with SNPs or UTRs. The software pipeline can also be used to obtain multiple independent RNAi designs per gene for independent validation of RNAi phenotypes. Additional strengths of NEXT-RNAi are its speed in designing comprehensive libraries and the generation of HTML reports including a variety of output options.
RNAi screening is being used increasingly in diverse organisms that only recently became amenable to genomic approaches. NEXT-RNAi can be deployed to design RNAi reagents for any sequenced genome to facilitate a better understanding of gene function through improved RNAi tools. This can be of particular utility for emerging model organisms that are suitable for large-scale RNAi studies but lack RNAi libraries. Further, in contrast to various microarray platforms, little attention has been paid to the re-annotation of existing RNAi screening data. We provide a fast and flexible software that accelerates the construction of consistent phenotypic data sets from RNAi screening experiments and helps to functionally annotate genome sequences.
NEXT-RNAi requires a defined set of files and parameters as inputs. Sequence input files are provided in FASTA format; feature input files, such as transcript-gene relationships or the locations of SNPs and UTRs, are provided in a tab-delimited format using defined names in the header row. Genome annotations and sequences for Drosophila were obtained from FlyBase ; Tribolium annotations and sequences were downloaded from BeetleBase ; Anopheles annotations and sequences were downloaded from VectorBase ; and all annotations and sequences for the human genome were obtained from the NCBI RefSeq database .
NEXT-RNAi is implemented in Perl. It requires the installation of Bowtie  and Primer3 . To utilize all options of NEXT-RNAi, the BLAST , BLAT , RNAfold  and mdust  programs are also required. On a Linux server (two Intel Xeon Quad-core 2.00 GHz CPUs, 16 GB RAM) running Ubuntu 9.10 server edition, the design of a genome-wide RNAi library for the Drosophila genome with approximately 70,000 constructs took about 4 hours. NEXT-RNAi software, installation packages and instructions for Linux and Mac operation systems and further documentations are accessible via . In addition, a platform-independent virtual machine (running on VirtualBox) with NEXT-RNAi and all dependencies pre-installed is available for download. NEXT-RNAi is used as a command line utility with parameters provided in an options file that allows specification of the design and annotation parameters (Additional file 11). An interactive mode that prompts for all necessary settings has been implemented.
D.Mel-2 cells (Invitrogen, Carlsbad, CA, USA) were grown in Express Five SFM (Invitrogen) supplemented with 20 mM Glutamax I, 100 U/ml penicillin, 100 μg/ml streptomycin. Total RNA was extracted using Trizol (Invitrogen), followed by Rneasy cleanup (Qiagen, Hilden, Germany), including on-column DNAse digest. mRNA was isolated with the MicroPoly(A)Purist kit (Ambion, Austin, TX, USA) and the RNAseq library was prepared according to Illumina's mRNA Sequencing Sample Preparation Guide. Paired-end reads were aligned to the D. melanogaster genome using Tophat  and RPKM values for each gene calculated with Cufflinks  based on the D. melanogaster gene annotation release 5.13 obtained from Ensembl. The data have been deposited in NCBI's GEO and is accessible through GEO Series accession number [GEO:GSE21283].
Long dsRNAs were synthesized using a two-step PCR procedure followed by in vitro transcription as described in . The concentration of each dsRNA was determined by photospectrometry and normalized to 50 ng/μl. We aliquoted 250 ng of each reagent in 384-well plates, and D.Mel-2 cells were added to the plates for an incubation time of 5 days. mRNA knock-down was measured by quantitative real-time PCR of two biological replicates using a SybrGreen assay (quantitative real-time PCR primers were designed using QuantPrime ).
The companion website to NEXT-RNAi at  contains extensive documentation and enables downloading of the complete software. The website also hosts complete NEXT-RNAi outputs for all pre-designed libraries, library evaluations and other analysis done for this manuscript.
We are grateful to Amy Kiger, Wolfgang Huber and Robert Gentleman for helpful discussions. We thank Stephanie Mohr and Norbert Perrimon for providing DRSC library information. TH is supported by a PhD fellowship by the Studienstiftung. TS is a postdoctoral fellow of the CellNetworks Cluster of Excellence [EXC81]. This work was in part supported by funding from the Deutsche Forschungsgemeinschaft, the Human Frontiers Sciences Program, the Helmholtz Association and the European Community's Seventh Framework Programme FP7/2007-2013 under grant agreement n° 201666.
TH and MB developed the concept. TH wrote the software and performed all calculations presented in the manuscript. TH and TS carried out the experimental validation of RNAi reagents. TH and MB wrote the manuscript.


This one had a lot of under-the-hood work to further the architecture goal of building the "Major" factions (human player, The AI) with mostly the same toolbox used for the "Minor" factions (Dyson, Devourer, Nanocaust, etc). This implicitly makes them all much more moddable and flexible.
But the result that stands out is that the Special Forces mechanic now stands out. Or rather, it used to be a mechanic, but now it's a faction in its own right. The main AI makes donations to the SF, but the SF decides what to build and where to go. As in AIWC it acts to defend the AI's planets against attack. Unlike AIWC, this behavior is more emergent (it should defend the AI against attacks from factions other than you, for instance) and is entirely moddable. If you want to make a SF sub-type that only builds long-range ships and only does sneaky drive-by attacks against the player, that can be done by modding. Or if you want a sub-type that only builds Etherjet Tractors and Widow Guardians and tries to tow the player's fleet to its destruction, that can be done too.
Any word on if/when AIW2 will be on GOG?
They want to see a build first, but we haven't been ready to share one with them yet. Based on our past history and conversations, I imagine that the game will launch on GOG at the same time it does Steam. But I can't really promise anything, since that's mostly out of my hands. At the moment they've been very interested in playing it, but I'm not keen on giving them an in-progress beta since they've turned down some of our past stuff.
Sounds good! Hoping for the best!


text:
Forest of Shadows é uma banda sueca de doom metal.

Biografia
Forest of Shadows nasceu na cidade sueca de Halmstad, em 1997. Inicialmente, Niclas Frohagen montou a banda como um projecto a solo, mas após o lançamento da demo The Silent Cry em 1998, as coisas tornaram-se mais sérias.

O multi-instrumentista Micce Andersson juntou-se a Niclas e a banda mudou-se para Gotemburgo. Pouco depois, Martin Claesson entrou no projecto, e a banda começou a dar concertos.

Em 2000, a banda começa a escrever aquilo que será o mCD Where Dreams Turn to Dust. Antes da gravação deste álbum, Martin Claesson deixa a banda. Mais tarde, a gravadora que iria lançar Where Dreams Turn to Dust abre falência.

A gravadora Rage of Achilles oferece-se para lançar o mCD (Outubro de 2001).

No Verão de 2002, Niclas mudou-se para Estocolmo e a banda voltou a ser um projecto a solo. Mal Niclas acabou a produção do primeiro album, Rage of Achilles cancela a edição devido a problemas financeiros. E mais uma vez, aparece uma gravadora disposta a comercializar o álbum, a Firebox. Em Outubro de 2004, Departure é apresentado ao público.

No ano seguinte, Where Dreams Turn to Dust é re-lançado.

Membros

Actuais
 Niclas Frohagen – guitarra, vocais, baixo, bateria

Discografia
 1998 – The Silent Cry (demo)
 2001 - Where Dreams Turn to Dust
 2004 - Departure
 2005 - Where Dreams Turn to Dust (re-lançado)
 2018 - Among The Dormant Watchers

Ligações externas

Bandas da Suécia
Bandas de doom metal da Suécia
Bandas de metal gótico
Bandas formadas em 1997

redpajama_set_name: RedPajamaWikipedia


Carol to Care is our annual Holiday event which consists of singing Christmas Carols in order to raise funds and awareness for homeless youth. All funds go towards youth shelters in our local communities. No singing experience is required! Attendees can receive community service hours for attending the event.


Private 3 bedroom 2.5 bath town home close to shopping, dining, roswell historic district, chattahoochee nature center and GA 400. New windows throughout, tile, luxury vinyl plank, and new paint. Two car carport with additional storage and fenced in patio. This one won't last long.
Listing provided courtesy of Ashman Associates, Inc..


This law explains why rivers, trees, snowflakes and even our vascular system exhibit the same branching design. It is also why hierarchies persist in human systems. For top-down flow, a hierarchy, is the optimal design for flow. Other than in nature though, human systems tend not to have the freedom to create fluid, evolving hierarchies.
So why is this important or interesting? Flow is a key notion when seeking to enable strategic agility. In the world of Lean manufacturing, optimising flow has always been central and it is becoming increasingly central in the Agile world as well.
I found it useful to make sense of this idea of flow using the Cynefin framework (please note, this is my own working hypothesis – a half-baked potato if you will, so it is not presented as gospel …).
I am fully aware these aren't all the flows (i.e. it's not complete) and the primary flows in obvious also exist in complicated and all of them exist in complex as well. I.e. work, resources, information, knowledge and many others also flow in complex to which is added the idea of fluidity. So read this as the primary flows we focus on in each domain, but not as exclusive categories.
Considering only the 4 primary domains, it makes sense for me to think about is as follows: In Obvious contexts, completely predictable, repetiive routine e.g. manufacturing contexts, the primary flows are flow of work and resources. In complicated contexts, where we're dealing with known unknowns, but where cause and effect relationships are not obvious and analysis and expertise are required, in addition to work and resource flows, we need to focus on the effective flow of information and knowledge. These two ordered system domains exhibit linear cause and effect relationships and stable contexts, hence the focus is on efficiency and optimisation.
In the chaotic domain, it was a challenge for me to think of what needs to flow here. A conversation at the conference led to the idea that maybe the flow that matters here is that of energy i.e. to contain the energy (emotional and otherwise) released in the chaos and ensure it flows in productive ways, not in desctructive fight/flight/freeze responses.
It is in the complex domain where it prof Bejan's work becomes particularly interesting. My hypothesis is that in complexity, in addition to flow of work, resource and information, systems need FLUIDITY, i.e. the ability to adapt our flow processes while in the flow. In the complex domain, the context is not stable: this is the so-called VUCA world. It is therefore critical for the system to be able to morph it's structure.
So then the question is how to enable fluidity. This seems to be a key aspect of both Prof Bejan and Prof Cilliers thinking. Hierarchies must be fluid; flow systems need freedom to morph their design. Simply focusing on introducing Lead or Agile methods at a team (or even organisational) level will not produce this fluidity.
If one deconstructs the idea of agility, it seems to require three things (at least) … mindsets; practices and skills; enabling structures. In most so-called Agile transformations the focus is mostly on process i.e training people in skills and practices. We neglect the mindset shifts and enabling structures that are needed to facilitate a cultural shift towards agility. Most people who've been through Agile training still lack a basic understanding of what agility actually is; why it's important; and what principles the recipes they were taught is based on. They most certainly are not more comfortable with uncertainty and ambiguity and while incentive and reporting structures remain unchanged, they are not incentivised to truly collaborate and act in the best interest of the collective.
Fluidity requires multi-directional flow, not the predominantly one-way flow that hierarchies are optimised for. This raises the question if the current shift towards networks and structures as described in among others Stanley McChrystal's Team of Teams is evidence of social flow systems morphing their design to optimise for the new kinds of flows they need to survive?
In nature, form follows design. In human systems we tend to design first and then try to force flow through our designs. I believe that we need to focus on creating fluidity, not only flow and this will require adaptive, emergent organisation designs that are fit for each unique context.
ensure organisational structures and processes are congruent with achieving fluidity i.e. don't create cognitive dissonance by stating one priority but enforcing the opposite through performance targets and other processes and sytems.
The idea of flow captures a lot of useful features of the relationships between internal and external features of a system.
Ideas on what needs to be done to promote adaptive ways of working seem to have stabilised in the discussion and literature as far as it relates to relaxing control and enabling emergence.
The gap in my mind is how we foster coherence and maintain a sense of purpose, to counter the characterisation of agile working satirised in the Dilbert cartoons as "… no more planning and no more documentation, just start writing code and complaining".
That seems to me to be the tension about which little is written – how to enable an adaptive organisation while ensuring some level of alignment and purposeful evolution as opposed to reactive adaptation.


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Rents for office space likely to rise: Report
OFFICE landlords seem to have enjoyed a recovery in this quarter with higher rents on the cards, according to a new report yesterday.
The demand for space is coming from small- and medium-sized enterprises (SMEs), mainly in non-financial industries such as media, information and communications, and businesses in professional and business services, said DTZ Research.
Another sign of a more buoyant market is seen in the net absorption rate, which increased to about 350,000 sq ft from July 1 to this point in September.
This is double the net absorption recorded for the three months to June 30.
Net absorption is the difference between the amount of space placed on the market and the amount leased.
However, while occupancy rates have increased in most areas due to the greater take-up during this period, the islandwide occupancy rate fell by 1.6 percentage points to 94.6 per cent compared with the three months to June 30.
DTZ said that this was due to the completed Asia Square Tower 2, Metropolis Tower 1 and Nexus@one-north. These buildings added more than 1.4 million sq ft of vacant office stock in this quarter.
Ms Cheng Siow Ying, DTZ's executive director of business space, said: "Besides the expansion of space by existing occupiers within the Central Business District (CBD), take-up of office space in this period primarily stemmed from small- to medium-sized firms."
Ms Cheng added that the smaller space requirements by these firms meant that "demand figures have remained relatively modest in 2013".
DTZ also noted that more service office operators have been catering to these firms by increasing the available space.
For example, CityHub committed to a new space in Tung Centre in Collyer Quay, and Regus will open offices in the JTC Summit building in Jurong East.
Ms Cheng added: "Nevertheless, the slow and steady build-up of demand has resulted in a tighter leasing environment where landlords are now less flexible on rental negotiations and instead have been holding or increasing their asking rents."
Higher rentals were seen in most areas, particularly in the newer and better quality buildings, said DTZ.
For example, average gross monthly rents in Raffles Place in the CBD are around $9.40 per sq ft this quarter, up 1.3 per cent on the second quarter, according to DTZ.
Rates in the Shenton Way, Robinson Road and Cecil Street area rose 3.4 per cent this quarter to $7.50 per sq ft a month over the previous quarter.
DTZ's Singapore research head, Ms Lee Lay Keng, noted that office space demand will remain modest, "as cost-conscious occupiers still prefer to renew existing leases".
She added: "Average rents in the CBD are anticipated to rise on the back of strong occupancy rates and an expected economic improvement, but this increase will be gradual."
Source: Straits Times, 25 Sep 2013
Newer Post2 floors at GB Bldg on sale for $3,673 psf
Older PostUnited Engineers buys two HP buildings for $402m

redpajama_set_name: RedPajamaCommonCrawl


Kessen II is a terrific game, has a really nice story plot and excellent game play. The graphics are exceptional the cut scences are realistic and beautiful. this game has passed Kessen 1 greatly. A must buy game for anyone who loves war games. Try to play Kessen one though since it would probably get you used to the whole thing and the confusion of some parts.
The whole story is fairly short only like four battles.
Graphics 10/10 : Like you've before this is also very important. The Graphics made my jaw dropped to the floor, Computer animated characters are amazing. Detailed Graphics from the characters armour and sword to the wavy smoke remains of a cannonade. I personal like seeing the soldiers go flying in the air going AH screaming! its funny.
Sound 7/0: The music of the game is pretty cool too. Japanese music mostly, when in battle it has the same music over and over and over... still it's very entertaining just to sit and listen. Some of the music can be very annoying and just plain annoying I don't know just listen and see.
Replay Value 9/10: I've played this game a few times, after you beat the game with the good side you can see how it was fighting for the side of evil. Seeing both endings is great. I personal Enjoyed the Evil ending heheheheheh.
Buy or Rent: Buy for sure, when your done you could always trade it in for something better. Try the First Kessen first. I would also reccomend the third one once it comes out sounds like an interesting game. Seriously though buy this game and play it you'll enjoy it.


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Explore state by state cost analysis of US colleges in an interactive article
Lamaip/iStock/GettyImages
Participle vs. Preposition
Christopher Cascio - Updated May 10, 2019
The Difference Between Participles & Verbs
Participles and prepositions are the source of confusion for many who wish to better understand the mechanics of English grammar. Both elements of the language are used in phrases, and both function as modifiers. Notwithstanding, participles and prepositions -- for the most part -- play separate roles in the architecture of a sentence as they are separate parts of speech; prepositions are one part of speech, and participles are verb forms, otherwise known as verbals.
According to the Purdue Online Writing Lab, a participle is defined as a verb "that is used as an adjective and most often ends in -ing or -ed." Because you use participles as adjectives, they modify nouns and pronouns. Participles come in two forms: present participles and past participles. Present participles end with -ing and are often confused with gerunds. Past participles end with either -ed, -d, -t, -n, -en or -ne. For the sake of clarity, when using participles you should place them "as close to the nouns or pronouns they modify as possible."
The Writing Center at the University of Ottawa defines a preposition as a word that "links nouns, pronouns and phrases to other words in a sentence." They often signal spatial relationships between the words they objectify and the rest of the sentence. Common prepositions of this type are "in," "on," "at," "above" and "beneath." Prepositions can also indicate time-related affiliations, such as "during," "before," "after" and "until."
Participial Phrases and Prepositional Phrases
Both participles and prepositions function as parts of grammatical units called phrases. Participial phrases include the participle and the word it modifies, along with any noun phrases or pronouns "that function as the direct object(s), indirect object(s), or complement(s) of the action or state expressed in the participle," Purdue explains. Participial phrases, like participles by themselves, always function as adjectives. A prepositional phrase consists of the preposition, its object (or the word it presents) along with any correlative adjectives or adverbs. Prepositional phrases, unlike participial phrases, can serve as adjectives and adverbs.
Participial Prepositions
Aside from the fact that both parts of speech -- participles are verbals -- are used in phrases that function as modifiers, another point of confusion arises from the overlap, which comes in the form of the participial that plays the role of a preposition: "Provided the meeting starts on time, it should still be light out by the end." Here, "provided" is the participle. Grammarist.com ascribes the term "participial preposition" to this usage, and claims these participles have become accepted as prepositions because of "widespread, long-time usage habits," when they would otherwise be considered dangling modifiers. Common examples include "assuming," "barring," "considering," "during," "given," "notwithstanding," "provided," "regarding" and "respected."
What Is a Present Progressive Verb?
How to Find the Parts of Speech in a Sentence
Difference Between Nominative & Objective Pronouns
What Are the Jobs Nouns Do in a Sentence?
Can a Prepositional Phrase Stand Alone as a Sentence?
Can a Participial Phrase Be in the Middle of a Sentence?
How to Tell the Difference Between Singular and Plural Verbs
How to Distinguish Between Adverb & Direct Object Infinitives
Purdue Online Writing Lab: Participles
University of Ottawa: The Writing Centre: What is a Preposition?
Grammarist: Participial Prepositions
Christopher Cascio is a memoirist and holds a Master of Fine Arts in creative writing and literature from Southampton Arts at Stony Brook Southampton, and a Bachelor of Arts in English with an emphasis in the rhetoric of fiction from Pennsylvania State University. His literary work has appeared in "The Southampton Review," "Feathertale," "Kalliope" and "The Rose and Thorn Journal."
Copyright 2021 Leaf Group Ltd. / Leaf Group Education


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Features Gantry 5 Addons Download
790 artisans receive training in KZN
A total of 790 artisans in various fields are receiving training in KwaZulu-Natal as part of the Human Resource Development Council (HRDC) Secretariat programme.
This was revealed on Friday at the Human Resource Development Provincial Coordination Forum meeting which was held at the King Shaka International Airport, in Durban.
The Forum is a structure of Council established by the Human Resource Development Technical Working Group as a forum for the provinces to ensure coordination, alignment, integration, communication, collaboration, implementation, and reporting of HRD imperatives with the Provincial Growth and Development Plans (PGDP) as well as Local Economic Development Plans.
KwaZulu-Natal as the host province shared achievements and challenges from the last quarter of 2014/15 financial year.
HRDC Secretariat Lineo Ramataboe said the province has awarded 2 043 external bursaries to out of school youth focusing on scarce and critical skills and 798 bursaries were awarded to students to study in Cuba.
There has been an increase of Academic Staff with Masters from 79 in 2012 to 109 in 2014.
A total of 1 852 public servants have been trained and 1 265 bursaries have been awarded to staff.
"In line with the National Development Plan to produce 30 000 artisans per annum, by 2030, the meeting also received a presentation from the Air Traffic Navigation Services (ATNS) on how provinces are taking full-advantage of bursaries for training as artisans they offer, to young people," Ramataboe said.
She said the Department of Science and Technology made a presentation on progress achieved in the development of a Research Information Management System (RIMS), that seeks to create synergy on data-collection for Research and Development.
"The system will provide reliable and comparable data for national surveys as well as routine information required by different statutory bodies.
"The data available can assist decision makers to craft informed policies based on reliable information," Ramataboe said.
Last modified on Monday, 24 August 2015 13:25
Lineo Ramataboe
HRDC
PGDP
Local Economic Development Plans
ATNS
Department of Science and Technology
NSFAS receives 10 000 applications for free education bursaries on day one
20 million rand for Mpumalanga education
Public Works helps 30 learners to study further
SA students to participate in supercomputing competition in Germany
Youth-led solutions for SA energy challenges
More in this category: « Learners encouraged to consider artisanship 38 people trained on conflict resolution »
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Feather River Route Capsule History
The Western Pacific Railroad was built in the last decade of the nineteenth century and the first decade of the twentieth century, to form the Pacific outlet for Jay Gould's Missouri Pacific and Denver & Rio Grande Western railroads. In later years, after Gould interests had lost control, and after D&RGW had secured direct western access to Denver, WP formed an alliance with D&RGW and Chicago, Burlington & Quincy to run trains between Oakland and Chicago. In 1949, this led to the introduction of the famous California Zephyr streamliner, with its multiple dome cars. In 1957, part of the line in the lower Feather River region was relocated by the builders of the giant Oroville Dam, to get it out of the area to be flooded when the dam was completed. CZ was taken off in 1968. (Amtrak later revived the name, but Amtrak's train uses the ex-SP Donner Pass route, not the Feather River route). In 1982, WP disappeared into the maw of Union Pacific, in a merger that also saw Missouri Pacific incorporated into UP.
The "high-line" to the GN connection at Bieber was constructed in the early 1930s.


Very well made and did fit very good.
Reviewer: Alexander from Lyubertsy, MOS. OBL.
Haven't used yet but it looks great.
Very pretty, I want another set.
Steel/Titanium is good and well made. There are not enough spacers delivered with it and I find the plastic knods to weak but until now I had not the chance to drive around.


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Home / News / APC Chairman, Adams Oshiomhole makes terrible English blunder (Video)
APC Chairman, Adams Oshiomhole makes terrible English blunder (Video)
Naija Broads August 10, 2018 - News
The Chairman of the All Progressives Congress (APC), Adams Oshiomhole has been captured in a trending video making a terrible blunder of the English Language while addressing the press.
Recall Senator Dino Melaye slammed the National Chairman of the APC, Adams Oshiomhole for claiming that the seat of the Senate President belongs to the ruling party.
Melaye also accused Oshiomhole of exhibiting what he called "brain arthritis."
The lawmaker added that the Senate presidency seat belongs to the Federal Republic of Nigeria, rather than to the ruling All Progressives Congress.
Melaye pointed out that the opposition Peoples Democratic Party gave the block vote that made Senator Bukola Saraki emerge as President of the Senate.
The lawmaker, who is one of the 14 senators that have so far defected from APC to PDP, said this while briefing journalists in Abuja on Sunday.
Oshiomhole had asked Saraki to resign as Senate President since the lawmaker had defected from the ruling to the opposition party. According to the APC chairman, the seat belongs to the majority party.
Melaye, however, condemned Oshiomhole for referring to the seat of the Senate President as a 'crown' that belongs to the APC. According to him, the seat is national and not political.
He said, "For those who are saying that Saraki should resign (as Senate President), I am so disappointed in the National Chairman of APC.
"He was very busy with labour and social issues that he did not take time to look at the Constitution and our extant laws to know that crowns are worn in the kingdoms, villages and communities; that there is no crown in the National Assembly.
"He has appropriated the seat of the Senate President of the Federal Republic of Nigeria to a political party. That is intellectual stagnancy. He has exhibited what can only be as a result of brain arthritis.
"I cannot understand why the national chairman of a party will not know that the seat of the Senate president is of the Federal Republic of Nigeria and has no party affiliation. Then, he appropriated the seat to APC, forgetting that even those who elected the Senate President were from the PDP.
"The PDP gave the Senate President 42 votes out of the 53 votes that he got on the day of election. So, the APC is the minority, even in the election of the President of the Senate.
"But the National Chairman of the APC, in his 'bolekaja' approach to life, myopically, intellectually stagnantly… anyway, you can't give what you don't have."
APC Chairman, Adams Oshiomhole makes terrible English blunder (Video) Reviewed by Naija Broads on August 10, 2018 Rating: 5


3D airplane flying game for cell phones to play online. Play as an airplane pilot and fly through a beautiful 3D world. Collect gems and stars during the way for higher score. Avoid crashing into clouds and ground.


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LGBT Marches Across USA Share The One Message The World Needs Right Now - Of Love & Togetherness
Susmita MukherjeeUpdated on Jun 14, 2016, 12:22 IST
The massacre of Orlando's Pulse nightclub led to a strong statement being made at the 46th annual Los Angeles Gay Pride Parade, apart from messages from Washington DC and New York. Members held moments of silence for those who died in the violent attack. Instead of lashing out at haters, the community has maintained a silence for the victims with messages of love and strength.
The Orlando massacre is the deadliest mass shooting in American history.
Members showed solidarity in Boston, Massachusetts.
A parade marcher holds a sign in memory of the victims of the attack at the 46th annual Los Angeles Gay Pride Parade in West Hollywood, California.
Prayers and moments of silence were the key to most parades by the LGBT community across America after the attack.
Members also gathered at iconic spots of the LGBT community to send out messages of togetherness at such a sorrowful time.
Peaceful marches organised through America do make a remarkable statement of forgoing hate and violence.


The Adult Choir praises God in song at the 10:30 a.m. Sunday Mass. The Adult Choir is open to all parishioners high school-aged and above. Rehearsals are Thursdays at 7:00 p.m. from September through Pentecost.


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Lay Your Hands (Stargate Remix) - Simon Webbe - Lay Your Hands (CD)
30.11.2019 Mazuzshura Comments: 9
Label: Innocent - 094633211308 • Format: CD Maxi-Single, Enhanced • Country: Europe • Genre: Pop • Style: Ballad, Vocal
I Love It When It Rains (Skit) - Keith Murray - Rap-Murr-Phobia (The Fear Of Real Hip-Hop) (CD, Albu, Dulce Matias - Mel D'cana (CD, Album), Black Power - The R.I.S.E. Project - Maroon (CD, Album), Euro-Trash Girl - Cracker - Low (Vinyl), Kinky / Freaky - RuPaul - Red Hot (File, Album), What Goes Up - Spaceant - Boomin - Wicked Body Movers - Vol. 2 (CD), Tek Tha Supah Latin - Daddys Home (CDr, Album), Wanna Be Gs, Du Hast Mich, Du Hast Meine Liebe, Focus (Remix) (Instrumental) - Joe Budden - Focus (Remix) (Vinyl), African Scaredy Katz In A One-Exit Maze - Sister Souljah - 360 Degrees Of Power (CD, Album)
Jan 01, · Check out Lay Your Hands (StarGate Remix) by Simon Webbe on Amazon Music. Stream ad-free or purchase CD's and MP3s now on abebalrepphohan.techteningluakarlumarrifusupatribb.co
Yobei says:
"Lay Your Hands" is the first single released from Blue band member Simon Webbe's debut solo album, Sanctuary. The single peaked at #4 on the UK Singles Chart, also faring well Genre: Pop.
Mashura says:
Label: Innocent - SINCD76,Innocent - • Format: CD Single • Country: UK • Genre: Pop • Style: Ballad, Vocal Simon Webbe - Lay Your Hands (, CD) | Discogs Explore.
Kitaur says:
Simon Webbe Lay Your Hands Simon Webbe - Lay Your Hands  (CD, Single) Innocent, Innocent.
Kedal says:
Lay Your Hands. Sometimes life can be a burden, Trying to stay one step a head, Feel the world up on my shoulder each time, I'm standing out on the edge, And my hopes have all deserted me, Like they've washed away in the sand, And it's hurting my pride, trying to survive, But I know that I stand a chance, When you lay your hands, Cos it's the only thing I have that still makes sense, Oh, baby.
Doulmaran says:
Sep 02, · TheLyricMasterz does not own the copyrights for any of the songs used, we take no credit whatesoever and have no affiliation with the artists/record labels o.
Maugis says:
"Lay Your Hands" () "No Worries" () "After All This Time" () "No Worries" is the second single released from Blue band member Simon Webbe's debut solo album, Sanctuary with backing vocals by Welsh pop star, Liam Kennedy.
Samumi says:
Sanctuary is the debut solo studio album released by Blue band-member and singer, Simon abebalrepphohan.techteningluakarlumarrifusupatribb.co album was released on November 14, , peaking at number seven on the UK Albums Chart, selling more than , copies in the United abebalrepphohan.techteningluakarlumarrifusupatribb.co album was later certified double platinum by the BPI. Three singles were released from the album: "Lay Your Hands", which peaked at number four .
Arashitaxe says:
Come and lay ur hands on me. Feel this road is getting longer now And i`m too far away from home Still I gotta keep on moving on But I can`t do it on my own Baby keep my head above water Help me swim for my life `Coz the game is getting harder The strain is gettin stronger And I can only face the fight When you lay your hands Oh yeah.


It all started in the Philippines, where I spent the first 17 years of my life.
I grew up in Apalit, Pampanga, a small town in a province north of Manila.
My parents were strict, enforcing a sole emphasis on education so my childhood consisted of 12 years at a private Catholic school where students were required to wear a uniform.
Books became my vehicle for transportation, taking me beyond the confines of school and my parents' rules. Dressing up became a form of expression, as I tired of the white collared blouses and navy blue pleated skirts we were required to wear daily.
A few days after I turned 17, my family moved to the Bay Area in California. The Bay nurtured many of my interests but it was only in the last year that I've finally been able to hone in on the two things that speak to me the most: fashion and literature.
Booklook, a play on the word lookbook, started as a way of capturing my internal temperature, which is almost always influenced by whatever book I'm reading.
Read books, serve looks. Join me.


Win the Race. With up to hp and outstanding fluid economy you don't have to decide between power and fluid economy, you get both. SEE POWER AND. The new 9RX 4-track tractors from John Deere provide superior power, performance, and ride quality when operating in the field. OLATHE, Kansas (August 25, ) – After years of design, field trials and customer feedback, John Deere introduces its first-ever 4-track tractor. Where Measured. Approximate. Horsepower. Model Name. John Deere Tractor Reference List. Canadian models. Commercial & Consumer Equipment division.


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package org.neo4j.ogm.annotation;

import java.lang.annotation.ElementType;
import java.lang.annotation.Retention;
import java.lang.annotation.RetentionPolicy;
import java.lang.annotation.Target;

/**
 * Annotation mainly focused to be used on the value attribute of another annotation.
 * It defines another attribute as its value that could be also represented by this one.
 *
 * @author Gerrit Meier
 */
@Target({ ElementType.METHOD })
@Retention(RetentionPolicy.RUNTIME)
public @interface ValueFor {
    String value();
}

redpajama_set_name: RedPajamaGithub


Holiday Hours - Have a great weekend!!
In spite of our studio being closed on Thursday, July 4th, you can still order a holiday arrangement for pick up or delivery today through Wednesday, July 3rd. Fresh flowers always add the perfect final detail to your 4th of July celebration!! Call us today to discuss your options (479.973.0588) and for our digital clients you can reach us via email at (althea@roseofsharonar.com).
And as always we love to see our clients at the studio -- 2015 C Main Dr, Fayetteville, AR.


Please login using your Toolkit username/email and password.
We welcome your feedback about the AUSRIVAS web site. To send us feedback, fill in this form and press the 'submit' button.
Please send us feedback about the AUSRIVAS web site. Is there anything missing, anything broken, or is there something you would like to see changed or added?


Growing up on the east coast, Paul mastered carving at a young age. Utilizing expertise in the backcountry, Paul began a career at Marmot Mountain Works, where he helped skiers from all over the world gather the best knowledge and equipment available. After Marmot's unfortunate closing, he found California Ski Company where he has become head bootfitter and backcountry master. He loves to spend his time getting back into the wilderness, climbing, hiking and, of course, skiing. These days if not bootfitting, he's usually cruising around Alpine or effortlessly ripping Mammoth.


Anyone here have a recommendation for a quality machine shop in North Orange county?
Doing a head gasket job for a friend and don't want to take the head to just anyone.
If you have a recommendation, let me know!
Looking for a shop in OC ??


You are here: Home » Criss Cross Mangosauce!
Criss Cross Mangosauce is a bilingual music and storytime created by two Latina moms who are performing artists and educators. The team brings their desire to foster children's love for languages, cultures and arts. Children of all ages and native tongues can attend this event at Mint Museum Randolph. Criss Cross Mangosauce is particularly catered for children from 2-8 years old, but everyone is welcome. 10:30-11:30 a.m. Free! Details.


One large ballroom with 50 meter floor-to-ceiling height and individual operable partitions that can divide the area into three separate ballrooms.
Ideal for meetings and training sessions for small to medium sized group.
Located at the topmost level of the hotel. Ideal party venue with a breath-taking view of the Taal Lake.
Designed as a modern conference venue. It is a Harvard style auditorium equipped with executive tables and chairs and state of the art avp facilities such as built-in projector screen, technical control room and individual outlets for laptops and electronic gadgets.


Access Mat Rental in Alberta & BC - WPW Inc.
WPW Inc. has an inventory of over 3500 access mats available for rent.
We provide hauling, setting, and removal of mats for both pipeline and facility applications.
Our mats are made of fir, which means they are lightweight yet durable. This allows for lower transportation costs, and is the reason why fir mats are becoming the mat of choice in our industry.
The use of access mats greatly reduces ground disturbance and is an environmentally friendly solution for pipeline right-of-ways and construction sites.
In addition to access mats we also have rig mats available for rent.


Smile and Wave: Best Family Weekend In Months!
Best Family Weekend In Months!
I wish I had a photo for every memorable moment from the last three plus days but other than Saturday morning at the pumpkin farm, we were either having fun at night or too into the moment to remember a camera! It was refreshing and just what we needed as a family to reconnect after so many weekends full of travel, Reserves drill, obligations, or deadlines with work.
On Friday after a trip to the grocery store and a run to grab some take out we decided on a whim (at 7:30 p.m.) to find a parking spot and have a picnic at Jordan Valley Park. It felt like those late summer nights in high school when the weather is perfectly warm and there are lots of people out enjoying the communal company of each other. The kids both got wet at the fountains and were out like a light when we finally put them to bed.
The next morning we made it to Anton's Coffee Shop and made friends with our waitress. It's still a toss up for us between Gailey's and Anton's for breakfast food but Anton's will be our go to if we're in a hurry. They have the greatest vintage high chairs made out of metal and covered in red and white gingham oil cloth and my omellete was delicious!
After breakfast we drove back to our neighborhood and spent a good hour at the pumpkin farm. Sebastian and Brett walked down rows of broccoli, climbed up piles of bark, and roamed through the pumpkins patiently stopping for photos at my request. Sebastian told me to take one of him on the wood chip pile and then made Brett do what he had done and took the photo below.
Later that evening was when we had invited our friends over to enjoy the fire pit so we got the house cleaned up a bit and decorated the front porch with pumpkins and mums. Our friends brought granny smith apples and caramel sauce and we had apple smoked turkey brats and S'mores with mint chocolate chip chocolate squares. We do weenie roasts extra fancy, ya'll.
The police showed up about thirty minutes after we'd started the fire and the rest of that story can be found in the previous two posts. It was even more sweet to have our friends there to enjoy our good news and get excited with us as we had been talking about it soon after they had arrived!
Sunday was spent enjoying our mostly clean house, playing with the kids, finishing projects, drinking coffee, fabric shopping, and making honey bran muffins among other things. I've gathered all of my supplies for pumpkin pie filled cupcakes and am prepared to make it work this week!
The leaves have just started turning here so it's starting to really feel like autumn despite the 85 degree weather. This week I'm excited to knock out some new sewing projects for my upcoming e-course as well as get some more handmade decorations up. Stay tuned for a fun tutorial!
Don't our kids look like they're having the time of their lives?
Hey that yellow big flower print fabric in the bottom image, i have that same fabric as a pillow case. I found it in an op shop here in Australia and used it to make a vest for my sister.
ps. I spent my weekend snuggled up inside with my husband watching movies as we had heaps of rain.
It was fantastic. Also uncovered a game we had forgotten we were given as a wedding gift, 'Monopoly City'. Its actually pretty rad and not a long as the original game.
twas a lovely one indeed.
This is such a great post! You sound so happy and the pictures are marvelous. I am so happy that the Vespa was recovered.
It looks like you had a lovely time! I love all those pictures you took!
aw, what a happy family picture! (well, I'm pretty sure the kids had a blast :) Happy Autumn!!
i love this post! such pretty pictures! it really looks like y'all had a good weekend and all this talk about fall and pumpkin patches really makes me miss the texas hill country.
it was a pretty nice weekend here in new orleans, though. a little warm for my taste this late in the year, but lots of sunshine!
There must have been something in the air because we had an equally wonderful weekend. :) Love all of the pics - fall is just the most amazing time of the year. We took the boys to IKEA on Saturday and yesterday was a trip to the orchard and the park. I love the memories we are creating with our children - they are really the best, aren't they?
adorable post :) such a lovely family.
upcoming e-course with sewing projects? sounds like fun!
love having magical weekends like that!!
love your sweet fam. we need a weekend like that. xo.
What a cute family photo at the end.
adorable photos! where is that pumpkin patch?! we took my niece & nephew to the wilson-rutledge farm on saturday... but that one looks cute!
I always forget the name but it's on Fort between Grand and Sunshine. Across from the big nursery, although they have pumpkins, too. It's a small family run business but the flowers are SOOO pretty!
Looks like a fun weekend! We went to our Church Picnic Saturday then Sunday night we went canoeing on a local lake. It was so beautiful! Wonderful way to end the weekend.


It's been two years since Mandy Harvey absolutely stunned the world with her viral "America's Got Talent" performance - a display that left the typically harsh Simon Cowell in awe. Harvey's vocals were on-point, but that wasn't the only reason Cowell, his fellow judges and the studio audience erupted in applause and a standing ovation.
The accolades came after Harvey revealed just before her performance that she's deaf, having lost her hearing about 10 years prior at the age of 18. It was a traumatic experience, as she could no longer hear the music she had grown to love.
"I lost the ability to hear the piano, and that's when I started to crack," she recently told "The Pure Flix Podcast."


text:
Q: Data change during doing Database backup I have a question regarding to mysql DB backup:
1.AT 8.00.00 AM ,I do backup Database by using mysqldump command.It somtime takes 5 second to finish.
2.While database backup is inprogress(At 8.00.01 AM ), someone make some changes to DB
Will backup version containt data changes of step 2 ?
I have goolge but not found explaination yet.Pls help me !

A: Percona provides a free tool for this purpose called xtrabackup.
For InnoDB tables MySQL uses log files to store DML commands, so that commands can be rolled back and some other things.
You can backup your database (in a non locking way), and after you created the backup you can apply the logs, so that you have a backup which holds the database status when the backup is finished. Don't know the commands without looking them up now, sorry, you'll have to have a look into the documentation.

A: It depends on your mysqldump command and which table the dump is on at the time of the update:

*

*If you used the --single-transaction option, then, no, it will not contain the late change.

*If you did not use that option then:

*

*if the table being updated has not been dumped yet, then yes it will have the change.

*If the table being updated has been dumped already, then no it won't have the change.

Chances are, you don't want that change in your backup, because you would like your backup to be consistent to a point in time.  Here is some more discussion about all those kinds of things:
How to obtain a correct dump using mysqldump and single-transaction when DDL is used at the same time?

redpajama_set_name: RedPajamaStackExchange


In investing, just as in life, risk is everywhere. Because the stock market is what the media follow so closely, you might have come to associate risk solely with the fluctuations in the price of equities.
But, risk is at the core of all investing. One way of defining "risk" is the possibility that the return you receive on any investment will turn out to be different than what you expected. Most of the focus, of course, is on lower than anticipated returns. Those are what make investing so scary to many.
That rate of return can be affected by many factors. And those possibilities will determine what options you choose for your 401(k). Now you have the option of using an advisor, the two of you can assess the pros and cons of each option. There are no wrong or right answers but an advisor will help you understand the risks and possible rewards of each. What ultimately matters is how much risk you should tolerate. Each person has a unique view of what is enough and what is too much. This measuring stick comes from their very unique life experiences. The really good advisor will coach the client to what risk is appropriate given the client age, income and other financial data. Then educate the client on the different kinds of risk. Here are a few to ponder. How each of these truly effect you will vary greatly!
This is the only one you hear about day in and day out. Market risk refers to the possibility that you will experience losses in your investment value. It is important to remember that the various market places are auctions. Because of this the ups and downs are driven by the perceptions of the bidders. This is a "systemic risk". Diversification itself cannot prevent the losses. But your portfolio might be configured with non-correlated assets to potentially minimize the effects of the ups and downs.
Whether it's a bond or a share of stock in a fund there is "business" or "unsystematic" risk associated with it. In the worst case scenario, the company could go bankrupt. That could be because of changing customer tastes, new technology, an upstart competitor, or litigation. Diversification helps cushion those kinds of losses. One way to diversify is through mutual funds. Most company retirement plans use funds.
It is a myth that bonds are "safe" investments. Those issuing the bond could default. Default happens when they stop making interest payments and can't repay the amount you have lent them. In investing the rate of return is correlated to relative level of risk. The corporate bond usually has a higher rate than a U.S. Treasury bond.
That advantage of municipal bonds is that they are frequently exempt from federal and/or state tax. For that they usually have a lower rate of interest than taxable investments. However, there is the risk that the rules could change mid-course and your bonds would be taxable. For example, the legislature could impose a tax on your bonds. That would lower your eventual rate of return. There is no additional tax advantage for a tax deferred retirement account so municipals would not likely be seen in your retirement plan.
With bonds, there is the possibility that the issuer will "call," that is, redeem the debt security before its maturity date. That can happen when interest rates are falling. In that environment, companies attempt to save money by issuing bonds with lower interest rates. That means you will have to search for an investment which provides the same return with the same level of risk. That might not be possible. When interest rates decline, the investor is usually forced to take on more risk in order to replace the same income stream. This is a form of reinvestment risk.
Inflationary risk is also known as purchasing power risk. Essentially it is the possibility that the value of an asset or flow of income will be eroded. How to manage this risk? The way is through choosing investments, such as stock funds which have a possibility of growth. That positions your portfolio to stay ahead of inflation over the long term.
Liquidity risk is associated with assets which cannot be sold quickly when funds are needed. The classic example is real estate. Liquid assets range from blue chip stocks to government securities.
However, you should also keep in mind that the U.S. Congress has the power to change laws affecting securities. That is specifically known as legislative risk.
In addition, society could pressure government to ban a product or service. If you own stock in companies in those sectors you could experience loss. That is social risk.
The bottom line on bonds is this: When interest rates rise their market value goes down. That doesn't affect you until you want to sell them. In the marketplace the value of your bonds will erode if the interest rates rise. That is because new issues of bonds will have a higher interest rate and be more attractive to a bidder.
Currency/exchange rate risk occurs when the price of one currency changes in relation to another. Suppose you have invested in a public company in Germany. The fluctuations in the currency in which that investment is denominated compared to the U.S. dollar could negatively impact the value of that security you own.
Yes, risk is at the core of investing. But it can be managed. A good financial advisor will educate you on risks and help you feel comfortable accepting the appropriate amount of risk. As an example a 30-year-old could likely completely avoid interest rate risk but take on some market risk. While an 80-year-old would likely accept some interest rate risk but be averse to too much market risk. Risk lives everywhere, understand it and risk can actually be your friend!
Whether you have a dollar or a million dollars in your retirement account, you'll be able to explore the value of a real advisor. Simply visit the Self Directed Brokerage Account advisor contact site.


My husband and I really enjoy entertaining at home.
I enjoy the preparations for our gatherings.
Planning new party menus, with dishes to dazzle.
Dressing the table, with my favorite pieces.
While our formal dining room is quite large, when we host guest, I often find myself wishing for just one more serving surface. With that in mind, I've been keeping my eye open for Home Bar Furniture. Moving my stemware to a bar would free up much needed space in my china cabinet, as well as provide the perfect place to hold our Bar Accessories for our home.
I am quite taken with the idea of a Folding Home Bar, and have my eye on a piece in Cherry Crosley.
The flexibility is wonderful, providing both storage and an expanding serving surface.
What more could a hostess ask for! Certainly, my in house bar keeping husband would love to show off his mixology skills, at this bar!
Having the right home entertaining pieces means I can spend less time juggling while serving my guests and more time enjoying them!
Tell me some of your favorite entertaining recipes or a funny story from your entertaining adventures! I'd love to hear them!
The opinions above are both honest and my own.
Bar images sourced from Cymax Stores.
This is a great looking piece. I would love to have one to!
I wonder if the 1st picture, comes in a darker cherry wood. It would look 100% better than what we currently use, when my husband play bartender.
Oh wow that's very pretty. I'd LOVE that in my home!


Plinka Design is a full service design studio.
We work with you to create a unique identity that conveys your brand message.
We design effective print materials, and take them from concept to delivery.
We create engaging websites with our emphasis on function, as well as form.
We design and produce impactful signage to further increase brand awareness.


KABUL (Pajhwok): the UN Assistance Mission in Afghanistan (UNAMA) in a statement on Thursday highlighted the need for elections free from fraud, and encouraged anyone seeing or suspecting fraud to file a complaint with the relevant Afghan authorities.
"All citizens should play an active, informed and constructive role in rooting out fraud. Afghans deserve elections that are inclusive, credible, transparent and free from any activity that could taint the results," said Tadamichi Yamamoto, the Secretary-General's Special Representative for Afghanistan.
The UN Mission notes that the Afghan electoral authorities have put in place numerous anti-fraud measures to detect fraud. The introduction, for example, of a voting system in which voters can only cast their ballot at a single polling station at which they are registered is arguably the most significant step taken towards mitigating fraud. The planned implementation of a biometric voter verification system should also contribute to deter and identify fraud.
Candidates also bear a major responsibility to prevent fraud and have an obligation to ensure that their supporters do not commit fraud in their name.
Any citizen suspecting or witnessing fraud has a civic obligation to file a complaint with the relevant Afghan authorities, especially the Electoral Complaints Commission (ECC ). As another new fraud mitigation measure, ECC officials will be present in every polling centre. Where credible allegations are made, Afghan law enforcement agencies should investigate expeditiously and, where appropriate, detain and prosecute those responsible.
The Mission also notes that the presence of up to 400,000 observers and candidates' agents, as well as political parties monitors, at the polling centres is crucial to ensure the transparency of the elections, and calls on observer organisations, political parties, and campaigns to play an active, informed and constructive role in safeguarding the integrity of the electoral process.
Afghanistan is slated to hold parliamentary elections on 20 October.


Please read Driving the North Road first!
To view photographs along the North Road, visit the Virtual Tour of the North Road. In this guide, the Km numbers are linked to these Virtual Tour pages. Please also see the Travelogues section of this website.
Beginning of the North Road, starting about 15 km north of Chicoutimi, off Hwy 167.
Km 11 Backroad to Chibougamau, logging road mainly, but not used by logging trucks every day.
Km 12 Waconichi Lake. There is a "Northern Arboretum" there, quite interesting to walk in the forest.
Km 20 The small Barlow River. CAUTION: The bridge here is very narrow and when there are logging trucks on the road, you might have to stop very quickly just before the bridge for them - this is dangerous if there are some more behind you!
Km 26 Backroad to Oujé-Bougoumou and Chapais -- some 46 Kms for Oujé, and 11 more for Chapais.
Km 32 Access road to Baie Penicouane, the southern part of Mistassini Lake. SEPAQ campground there, boat ramp, picnic tables.
Km 65 There is a spring on the left side (northbound) of the road here.
Km 77 Beaver Camp (road maintenance camp). Also access to the Lake & River Des Maures.
Canoeing: From here you can reach the Rupert River & Mistassini Lake by canoe, and then paddle down the Great Rupert! Or stay on the "South branch", the Natastan River until Lac de La Passe, portage your way through the Marteen River waters and paddle back to the road at Km 177. Or keep going over the Natastan River until the end, paddle all Lac Mesgouez and come out at Km 221.
Frotet Lake. This is the headwaters of the Broadback River.
Km 106 Cellular phones do work here for a short ways.
Km 108 Junction of access road for the Troilus Mine, 44 Kms away.
At about km 27 of this access road, there is a gravel pit, just over the hill after crossing the Broadback River. This is the access for "Square Tail Lodge" outfitting camps, a world class fishing spot, mainly for speckled trout.
The road turns west just after Km 108; there is a nice valley on the north side of the road. Lake Lezai is visible down there, quite pretty scenery with all the small islands in the lake.
Km 118 Here the road turns back north. Just at the curve, there is a side road on the left side to Cachisca & Opataca Lake. This road used to be another way to come up here, via Chapais & Oujé-Bougoumou, following the power lines for most of the way. In 1999 this side road road was completely washed out after the tremendous winter of 1999 (over 9 feet of snow!). When the Mistassini Lake ice melted in the spring (headwaters of the Rupert River), the Rupert River water level went up 5 feet and stayed like that until the summer of 2000! The water level was even higher in 2004, due to Mistassini Lake ice again. The ice went out only on June 20th of 2004. The water level was already quite high but it went up another 3 feet!
Cheniapiscau rest area. Picnic tables, toilets, scenic viewpoint, camping.
Camping & fishing note: This campsite and any other campsite before Km 164 are inside the SEPAQ territory. You need to register and pay for camping or any other activities inside the Reserve. No registration paper might mean a fine from the Quebec Game Wardens. They are all over the place in the summer. Same thing for fishing. First "free" fishing spot is the Marteen River at Km 178.
Km 154 Limit of the logging operations! Breathe a big sigh of relief (if you're traveling north)!
Here there is a road on the left side. Just past this (northbound), there is a steep hill. Stop here! Get out of the car, climb up the rocks on the left side (northbound) and have a look at the scenery! When the air is very clear, you can see all the way back to Km 108. And if you are familiar with the area, you can clearly see the Troilus Lake mountain, this is more than 60 Kms away!
Marteen River. By paddling downstream from here, you can reach the Rupert River after a week or so of paddling.
Km 221 Small side road on the right hand side (north bound): access for Mesgouez Lake & the Rupert River. There is an extensive set of Cree camps here (spring fishing camps). Boat ramp, camping, fishing.
Km 232 or so Southern limit of the WEE-SEES-INDOHOUN Game Management Territory. New in 2003. You will need a pass (free) to fish or camp anywhere inside this special zone. Pass can be obtained at the SEPAQ office at Mont Chalco, just about 5 Kms North of Chibougamau or at the Nemiscau Quebec Hydro camp (Km 290).
Rupert River Gorge. STOP HERE AND TAKE A LOOK.
Picnic tables (3 or 4), pit toilets (2), shelters (3), fishing. The rest stop is small and overused, and rather dirty.
Spectacular rapids!! The Rupert River isn't anywhere near as big here as it is downstream where it crosses the James Bay Road. But these rapids are a seething mass of wild roaring water nonetheless.
The Rupert River will be diverted, upstream from here, in late 2009. Get up here and see this magnificent wild river while it is still unaltered!
It is possible to camp here, but NOT at the rest area. Just before the bridge (Northbound), on the right side, there is a small road. This is the portage trail. There is room down there for 1-2 cars & tent.
The "cut-off" point for the Rupert River Diversion project is only about 4-5 kilometers upstream. At low water, it is possible to walk on the North shore of the River and reach this point, about a one day walk.
To have another view of the Rupert River, one can walk North on the road for +/- 300m -- there is a trail there. This is the portage trail, ends in a nice "pool", where there's a nice view of the second set of rapids (Class V).
There is also another trail. Just in front of the entrance for the small road (the portage trail mentioned above), at the south side of the bridge. This trail leads right on top of the chute, one can take breathtaking pictures from there. But take great care, it is almost scary!
And just past the east end of the bridge, on the north side, you can walk down to the river above the rapids.
Same note here about fishing: you are in the WEES-SEES-INDOHOUN zone, so you need a pass.
Km 241 Bras de Rupert - an arm of the Rupert River that is a lake.
Main Junction, Poste Albanel transformer station.
-Left turn for the North Road.
-Right turn for the Hydro Quebec Poste Albanel transformer station.
-Straight ahead is the road to "Camp Rupert".
-Straight ahead for the Hydro Quebec Poste Albanel transformer station.
-Left turn is the road to "Camp Rupert".
Lescar Hills rest stop. Picnic tables, toilets, scenic viewpoint (nice view to the north).
Hydro Quebec Nemiscau transformer station.
Camp Nemiscau, 0.5 km north of North Road.
Km 290.6 Relais Routier: fuel, meals, garage, information office. Gas bar open Monday-Saturday 6am-12noon, 1pm-7pm; Sunday 7am-12noon, 1-5pm. In May 2008 gas was $1.77/liter, diesel was $1.91/liter.
Km 290.8 Road to Camp Eastmain-1, Poste Nemiscau, Project Eastmain-1.
Access road to the village of Nemaska (Nemiscau), 10 km to the north. Gasoline, diesel, restaurant, store, lodging (819) 673-2512.
North of this point you will there is a lot less traffic, but the road is in poorer condition, with numerous soft patches. Also the speed limit drops to 70 km/h.
Going south, be prepared for a lot more traffic, but the road is in much better condition.
Km 354 Turnoff for Lake Nemiscau & Old Nemaska. The Cree village was located there until 1970 when the HBC closed the post and the people were "evacuated" by Hydro Quebec. "New Nemaska" is at Lake Champion (km 299).
Boisrobert Lake campsite and rest area. Picnic tables, toilets, boat launching ramp, camping.
Km 383 Extensive forest fire in 2007.
Km 385 Boisrobert Lake - sign is hidden in the bushes.
End of the North Road: junction with the James Bay Road at km 275. Pavement!


This is a list of battlefield map videos for Gettysburg, covering the fighting on the first day of the battle (Chapters 1-10), on July 1, 1863. These brief videos are intended as supplements for the book, Our Fathers at Gettysburg. You might find it helpful to view each just before reading that chapter, and/or again after you're read the chapter.


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Being grateful seems like such a natural and instinctive response, it can hard to believe that there's a science to it.
The results of this experiment showed that if you're going through "a particularly tough time" you're more likely to benefit from this exercise. That's got to be music to the ears of anyone going through a divorce!
Watch this video and then pick up a pen and write a letter, or call someone.
Guaranteed to change their day ... and yours!
Gratitude blocks toxic, negative emotions, such as envy, resentment, regret—emotions that can destroy our happiness.
Gratitude makes us look outside ourselves. Gratitude is an 'emotion complex' (like guilt or shame) that implies a connection with others. When we feel grateful, we're realising that outside forces – friends, families, the kindness of strangers, the 'universe' (or even a higher power if that's what you believe in) – are working to help us achieve good things in our lives.
Gratitude heightens our sense of self-worth. Think about it – if this complex network is working so hard to enhance your enjoyment of life, then they must really value you., which can change the way you see yourself.
Gratitude is motivating. Research shows grateful people take better care of themselves, exercise more regularly and are more helpful, generous and compassionate.
Gratitude is self-perpetuating. The more you notice, the happier you are, which makes you more open to noticing stuff to be grateful for.
There's a number of studies showing that in the face of serious trauma, adversity, and suffering if people have a grateful disposition, they'll recover more quickly. Perhaps it is because gratitude gives people a perspective, a basis of acceptance and a comparison, from which they can interpret negative life events and help them guard against post-traumatic stress and lasting anxiety.
Just because gratitude is good doesn't mean it's always easy. Practising gratitude can be at odds with some deeply ingrained psychological tendencies.
One is the "self-serving bias." That means that when good things happen to us, we say it's because of something we did, but when bad things happen, we blame other people or circumstances.
Gratitude also goes against our need to feel in control of our environment. Sometimes with gratitude, you just have to accept life as it is and be grateful for what you have.
Need some help to create the habit of gratitude in your daily life?
How can you benefit from counselling through the process of preparing to separate?
When should you leave your relationship?