Heterozygote and homozygote

[swbarnes2]

It's not an inconsistancy, exactly. Your problem is not that a bunch of reads are inaccurate. It's probably what the template really looks like; a mix of two different DNAs. It's almost certainly because the sample that was given to you was a mix of two slightly different populations. It happens, you should tell your submitters what you are seeing. You could confirm some with sanger sequencing, to be sure.

Hopefully, your mutant samples should be clonal, and not have mixed letters. What you'll want to do is check everything that looks like a possible SNP in your mutant, and see if the wt shows a trace of the mutant letter. If it does, than your mutant has the alternate letter because of descent, not becuase of mutation. The mutations you are most interested in are those where the wt doesn't show a trace of the alternate letter seen in the mutant.

[biocc]

thanks

I do bacteria, and I see mixed samples all the time, too.

The simplest explanation I can think of is that you've got a non-clonal population.

For instance, I've seen cases where I was given a non-clonal parental strain, and some offspring resistant strains, and there will be some mutations that are clean in the offspring, but mixed in the parental. I've tested some such loci with sanger sequencing, and they pretty much always confirm the Illumina data.

thanks for your reply. this question puzzled me for a long time. since I focus on the difference between mutated strain and WT, it may be difficulty to determine the SNPs from the mixed samples which you inferred. to verify the SNPs which BWA and samtoos gave, i visualized the alignment by Tablet, and found some reads support the SNP, some do not in almost of all SNPs. how to deal with this inconsistent? could you give me some advice ?

[swbarnes2]

I do bacteria, and I see mixed samples all the time, too.

The simplest explanation I can think of is that you've got a non-clonal population.

For instance, I've seen cases where I was given a non-clonal parental strain, and some offspring resistant strains, and there will be some mutations that are clean in the offspring, but mixed in the parental. I've tested some such loci with sanger sequencing, and they pretty much always confirm the Illumina data.

[biocc]

hi, when I resequenced a bacteria and aligned with reference , i found heterozygote is very common. since my sample is lab strain and bacteria is haploid， i do not understand this phenomenon. could someone explain for me? thanks