Understanding FREQ; Reasons for variant allele frequency to deviate from 50%

Question

[drabiza1]

What are the reasons to observe variant allele frequency (FREQ) generated on an Illumina DNA sequencing platform around 10-30% or 70-90% for heterozygous variants? As in what are the reasons that this value would deviate from 50%, assuming there is no aneuploidy. Is there any biological mechanism that can explain this or is this just a matter of PCR duplication during sample preparation.

Example: GT:GQ:SDP:DP:RD:AD:FREQ:PVAL:RBQ:ABQ:RDF:RDR:ADF:ADR 0/1:15:28:28:23:5:17.86%:2.5729E-2:32:33:18:5:5:0

Answer 1

[lethalfang]

Biologically, it can be copy number variations (e.g., https://pubmed.ncbi.nlm.nih.gov/17122850/). It also also be sequencing and bioinformatic biases, e.g., mismapped and/or misaligned reads.

Looking at your info RDF:RDR:ADF:ADR, seems to have 18 forward reference reads, 5 reverse reference reads, 5 forward variant reads, and 0 reverse reference reads. There seems to be a bit of strand bias, but that could be due to any reasons, e.g., if the variant position is on the edge of a capture region, etc.