Map to reference and MIRA assembly of mitogenomes in Geneious (Part 5)

In this last mitochondrial genome assembly tutorial we complete our assembly. We were not able to recover the control region, due to the high genetic distance to the reference genome. Also with de novo assembly with MIRA and mapping strategies with MITObim were not able to assemble the complete genome. However, the complete coding region was consistent with all assembly strategies, giving us high confidence that it was assembled correctly. The mitogenoe is published in the jornal Mitochondrial DNA:

Elbrecht & Leese (2015). The mitochondrial genome of the arizona snowfly Mesocapnia arizonensis (Plecoptera, Capniidae). Mitochondrial DNA

Download the MT sequence on GenBank http://www.ncbi.nlm.nih.gov/nuccore/?term=KP642637

Table of contents
0:01 Recap, problems when downloading the wrong SRA file for mitogenome assembly. The correct data set is this one here http://www.ncbi.nlm.nih.gov/sra/SRX450974
2:39 Identifying and obtaining the correct dataset using ncbi blast
4:58 Subset data and blast it against the reference genome
7:03 Taking look at the assembly. We did extract the wrong length of reads (99/101 while 100/100 is correct). Fixing that problem
8:46 Find the optimal reference genome using blast
10:14 Blast reads against reference genome and map to reference
11:08 Editing the mapped reads (fix errors in the contig)
13:45 Export the consensus sequence of the draft genome and map reads against the draft
16:17 Checking for improved coverage in second iteration
18:35 Why random subsetting of sequences is important
21:19 Assembly with 3 million sequences and both forward and reverse reads, editing flanking regions
23:28 Sequences should have been extracted with 100 bp length!
24:04 Assemble sequences against the edited draft, several iterations to close the control region (which did not work).
31:45 Assembly results with the program MITObim
33:00 De novo assembly of 1 million reads with MIRA (as a Geneious plugin)
33:15 Compare the results of the assembly strategies with an alignment of all consensus sequences and add annotations
26:29 Obtain a random sequence subset with randomFQ http://code.google.com/p/ea-utils/
28:32 Set paired end reads in Geneious (much better than exporting selections which we have done before)
40:45 Giving up on the control region, the rest gets assembly well with all 3 strategies!

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