I have assembled four bacterial genomes derived from MiSeq pair-ended sequencing data using the following steps:

1. Assembly using CLC Workbench;
2. Assembly using SPADES;
3. Assembly using A5 pipeline;
4. Merging of the three assembles using CISA;
5. Quality check of the assemblies using QUAST.

For checking the misassemblies, QUAST relies on a reference genome. However, for most of my draft genomes, I do not have a proper reference genome (too much genome differences in relation to those deposited in Genbank).

So, I ask you. How could I validate the genome assembly using intrinsic data? For example, using read mapping, what are the criteria to correct some regions? What is the best software for this purpose?

Thanks
I don't know any <em>ad hoc</em> solution. But you can try looking at:

 - fraction of reads that aligned - if many reads didn't aligned you probably miss some regions in your assembly
 - fraction of reads with concordant pairing (ie `samtools flagstat`) - if this is low, you have likely rearrangements or high genome fragmentation
 - pairwise genome alignments (ie. <em><a href="http://mummer.sourceforge.net/">nucmer</a></em> or <a href="http://last.cbrc.jp/"><em>lastal</em></a>) of your assemblies to check for large inconsistencies between them

It's always good to compare vs chromosomes of some relative species to check whether your assembly make sense.