Duplicating contigs in caf file

[Derek_S]

Hi all,

I'm wondering if anyone has experience in duplication of contigs for the final stitching of a denovo assembly of a bacterial genome?

I've done a 454/Illumina hybrid assembly in MIRA and am now trying to reduce the number of contigs by stitching contigs together manually. I have a decent reference genome (190 contigs for a 7Mbp genome) so I know where some of the 16S RNA repetitive regions should be to join some of the larger contigs. The problem I run into is when I join a non-repetitive contig to a repetitive one there is only one copy of the repetitive contig even though it should be represented in the genome 4-5 times (I'm using GAP5 to do this). Is there a way to duplicate these contigs so I can assemble them more than once and hence stitch the genome together better?

Thanks

Derek

[BaCh]

I'm wondering if anyone has experience in duplication of contigs for the final stitching of a denovo assembly of a bacterial genome?

You cannot immediately duplicate a whole contig as this would also duplicate all reads contained within ... and duplicate read names is something which will break MIRA, gap4, gap5 etc.

Two solutions:
a) you duplicate contigs completely, but change all names contained within
b) you duplicate contigs by just taking the consensus and insert these as single reads into the project.

I'd go the second way as the first involves a lot of work by hand. Doing this is quite easy:
1. use "convert_project -f caf -t fasta -s" to convert a CAF to single consensus FASTA files
2. take the consensus you want, rename the sequence to something you can distinguish (e.g. "RNAcopy_01") then use "convert_project -f fasta -t caf -m" to make a CAF.
3. Append the new CAF with RNAcopy_01 via "cat" to the complete project CAF
4. Rinse, repeat at step 2.

B