Question: High Mapping but No Reads Properly Paired
0
gravatar for pld
2.8 years ago by
pld4.8k
United States
pld4.8k wrote:

After mapping PE reads with bwa-mem, samtools flagstat gives me this output:

53681238 + 0 in total (QC-passed reads + QC-failed reads)
0 + 0 secondary
779896 + 0 supplementary
0 + 0 duplicates
48364704 + 0 mapped (90.10% : N/A)
52901342 + 0 paired in sequencing
26450671 + 0 read1
26450671 + 0 read2
0 + 0 properly paired (0.00% : N/A)
47584808 + 0 with itself and mate mapped
0 + 0 singletons (0.00% : N/A)
0 + 0 with mate mapped to a different chr
0 + 0 with mate mapped to a different chr (mapQ>=5)

Viewing this in IGV shows all reads mapping in FF or RR.

I've never seen something like this before, any suggestions?

bwa • 1.3k views
ADD COMMENTlink modified 2.7 years ago by Biostar ♦♦ 20 • written 2.8 years ago by pld4.8k

Any chance the data files are mislabeled? Do the headers look right?

ADD REPLYlink written 2.8 years ago by genomax65k

I don't think so. I'll double check.

ADD REPLYlink written 2.8 years ago by pld4.8k

did you reverse-complement one of your fastq files ?

ADD REPLYlink written 2.8 years ago by Pierre Lindenbaum119k

Not to my knowledge. Any chance this could have happened pulling the data off the machine?

ADD REPLYlink written 2.8 years ago by pld4.8k

No. It will have to be done consciously.

ADD REPLYlink written 2.8 years ago by genomax65k

Can you show us your bwa command line?

ADD REPLYlink written 2.8 years ago by Dan D6.7k
1

I'll just save myself the embarrassment: I supplied the forward read file twice.

I've been sitting here scratching my head and checking everything but the command I ran. Today is not my day.

ADD REPLYlink written 2.8 years ago by pld4.8k

Take comfort that each of us reading this thread will have a "not my day" day soon.

ADD REPLYlink written 2.8 years ago by Dan D6.7k

Hi! Can I resurrect this thread? I'm having the same problem, and I didn't supply the same file twice... Also, everything worked for the other populations I'm using, just one failed...

BWA command line: bwa mem -R '@RG\tID:pop1\tSM:PCN\tLB:library1' referenceP30 PCN_R1_30G.fastqsanger PCN_R2_30G.fastqsanger -a -t 16 -T 10 > PCN30.sam

ADD REPLYlink written 23 months ago by rohenriques0
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