Assemble mitochondrial genomes from short read data

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A simple procedure for assembling mitochondrial genomes based on whole-genome re-sequencing data. The first step is to extract reads from the sequence library based on a closely related entirely assembled genome (e.g., for pig, the MT genome as present in the genome build, but could also be of a related species). The genome is then assembled using SOAPdenovo.

  • a reference genome of a closely related population or species.
  • a bowtie2 index (make with bowtie2_build)
  • a blastable db of the reference mitochondrial genome
  • a SOAPdenovo configuration file:

soapdenovo.config 

 [LIB]
 avg_ins=450
 reverse_seq=0
 asm_flags=1
 rank=3
 q1=fq1.fq
 q2=fq2.fq

Note that the avg_ins flag may vary between libraries; may have an effect on assembly efficiency.

<source lang='bash'>

  1. !/bin/bash
  2. SBATCH --time=1000
  3. SBATCH --mem=16000
  4. SBATCH --ntasks=8
  5. SBATCH --nodes=1
  6. SBATCH --constraint=4gpercpu
  7. SBATCH --output=output_%j.txt
  8. SBATCH --error=error_output_%j.txt
  9. SBATCH --job-name=assemble_mito
  10. SBATCH --mail-type=ALL
  11. SBATCH --mail-user=hendrik-jan.megens@wur.nl

module load bowtie/2-2.2.1 SOAPdenovo2/r240 BLAST+/2.2.28 MUMmer/3.23

bowtie2 --phred$2 --local -p 8 -x mt_pig.fa -1 $3 -2 $4 | head -2 >$1_mito_align.sam bowtie2 --phred$2 --local -p 8 -x mt_pig.fa -1 $3 -2 $4 | awk '$5>0' | head -10000 >>$1_mito_align.sam

java7 -jar /cm/shared/apps/SHARED/picard-tools/picard-tools-1.109/SamToFastq.jar I=$1_mito_align.sam F=fq1.fq F2=fq2.fq INCLUDE_NON_PF_READS=True

SOAPdenovo-63mer all -K 63 -p 4 -s soapdenovo.config -o $1_mito_assembly.fa

blastn -query $1_mito_assembly.fa.scafSeq -db mt_pig.fa -outfmt 6

mummer -mum -b -c mt_pig.fa $1_mito_assembly.fa.scafSeq > mummer.mums mummerplot -postscript -p mummer mummer.mums </source>

Invoke like this: <source lang='bash'> sbatch do_mtalign_bowtie_pig.sh MA01F18 33\ 

/lustre/nobackup/WUR/ABGC/shared/Pig/ABGSA/ABGSA0071/ABGSA0071_MA01F18_R1.PF.fastq.gz\
/lustre/nobackup/WUR/ABGC/shared/Pig/ABGSA/ABGSA0071/ABGSA0071_MA01F18_R2.PF.fastq.gz
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