Definition: In this task, you're given a paragraph from the research paper and your task is to generate a suitable title for the research paper based on the given paper. Under 100 words is a good title length.
Input: was performed with RdRpSeq-Fwd (similar to the first round) and RdRpSeq-Rnest primer by using MyFi ™ Mix Kit (Bioline, USA) according to the following parameters: 95 °C for 1 min, followed by 35 cycles at 95 °C for 15 seconds, 56 °C for 15 seconds, 72 °C for 20 seconds, and 2 min at 72 °C for terminal elongation[5]. The resulted amplicons were separated by gel electrophoresis, and the DNA fragments were excised, purified and subjected to sequencing in both directions. The obtained sequences were verified by basic local alignment search tool. As for the amplification of complete N gene region, cDNA was synthesized by using SuperScript III First Strand Synthesis System for RT-PCR (Invitrogen, USA) with NR2 primer (Table 1). Two microliters of the generated cDNA was subjected to one round of PCR cycle by using MyFi ™ Mix Kit (Bioline, USA) with NF1 and NR2 (Table 1) primers following the parameters: 95 °C for 1 min followed by 25 cycles at 95 °C for 15 seconds, 52 °C for 15 seconds and 72 °C for 30 seconds and terminal elongation at 72 °C for 5 min. Then, 0.5 μL of the resulted PCR amplicon was further subjected to the second round PCR following the abovementioned parameters. The resulted PCR product was separated by gel electrophoresis while the DNA fragment was excised, purified and subjected to sequencing by using NF1, NR1, NF2, NR2 and N350R primers. The obtained sequences were analyzed by using ChromasPro version 1.7.4 (Technelysium Pty Ltd, Australia). Multiple sequence alignment was performed by using Clustal X (version 2.0.11) with 44 human-derived (including the Malaysian case; MyH010414; GenBank accession No: KP769415) and 3 camel-derived MERS-CoV sequences[6]. Maximum likelihood analysis was conducted by using Molecular Evolutionary Genetics Analysis 6.06[7]. The evolutionary history was computed by using Tamura-Nei model with 1 000 replicates of bootstrap analysis. Extracted RNA from combination of throat and nasal swabs from the case was detected positive both in rRT-PCR and conventional RT-PCR for partial RdRp and N gene. The cycle threshold values for upE, ORF1a and ORF1b were 28.19, 29.16 and 33.36, respectively. Amplification of partial RdRp and N gene regions resulted in DNA fragment of ~240 bp and ~310 bp, respectively (data not shown). Initial sequencing result of these fragments showed highly identical nucleotide sequence (99%) to MERS-CoV strain Jeddah_1_2013 (KJ556336.1), further confirming the MERS-CoV infection. JX869059/Human/Jeddah/HCoV EMC/2012 KC776174/Human/Jordan/Jordan N3/2012 Clade A 0.0005 67 63 63 63 66 50 62 Clade B Figure 1. Phylogenetic analysis of N gene region of Malaysia MERS-CoV case together with 3 camel-derived and 43 human-derived MERS-CoV sequences from 2012-2014. Red diamond: Malaysian case; Blank diamond: sequences from the camel-derived MERS-CoV.

Nucleocapsid gene analysis from an imported case of Middle East respiratory syndrome coronavirus, Malaysia Asian Pacific Journal of Tropical Disease