Anyone used BAMSurgeon to spike in 1KB insertions/deletions?

Question

[sichan]

Hi Everyone,

I'm interested in using BAMSurgeon to spike in structural variants up to 1KB in length to my BAM.

Here's my command:

python addsv.py -v SVs.bed -f Input_BAM -r hg19.fa -o Output_BAM > out.txt 2>error.txt

Where SVs.bed contains the following:

20    8189474    8190474    DEL    1.0
20    8771681    8772777    DEL    1.0
20    8823637    8823638    INS    myseq210.fa
20    11753526    11753527    INS    myseq731.fa

The last two entries are insertions and the myseq*.fa are fasta files and each contain a 1KB randomly generated nucleotide sequence.

Within my error.txt file, I see the following for each of the four SVs I'm attempting to spike-in:

encountered error in mutation spikein: 20    11753526    11753527    INS    myseq731.fa

Traceback (most recent call last):
  File "../addsv.py", line 435, in makemut
    maxcontig, refseq, alignstats, refstart, refend, qrystart, qryend, tgtstart, tgtend = trim_contig(mutid, chrom, start, end, maxcontig, reffile)
ValueError: need more than 2 values to unpack

My out.txt file ends with the following

INFO    2015-09-21 14:07:35.645967    no succesful [sic] mutations

So, none of the mutations were spiked-in successfully and no output BAM is created. I can share both files if necessary, but has anyone encountered these errors?

Also, if anyone knows of other tools to spike SNVs/Indels/SVs into BAMs, please let me know.

Thanks

[lzy]

Hi Sichan,

Could you tell me how get the correct BAM Alignment input file ? As I use the follow command lines:

1. bwa mem -t 2 -M hg19.fa NA12878_R1.fastq NA12878_R2.fastq > NA12878.sam
2. samtools view -bS NA12878.sam > NA12878.bam
3. java -jar SortSam.jar INPUT= NA12878.bam OUTPUT= NA12878.sort.bam SORT_ORDER=coordinate TMP_DIR=tmp_picard
4. java -jar AddOrReplaceReadGroups.jar I=NA12878.sort.bam O= NA12878_fixed.bam RGID=4 RGLB=lib1 RGPL=illumina RGPU=unit1 RGSM=20 (Then ValidateSamFile.jar shows No err found)
5. samtools index NA12878_fixed.bam

I do not know whether it is right. Do I need to re-align reads to the reference? If so, how to do this re-align ?

Thanks.

Answer 1

[Adam Ewing]

Hi Simon,

Could you double check that you have the latest from the repository (git pull) and see if you can get the test (test/test_sv.sh) to run?

Also, if you like, please send command lines / full error output to me ([e-mail]) and I'd be happy to have a look.

[sichan]

Thanks for your response, Adam. I just sent you an email.