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somatic CNV PoN

Dear all,

When I run the CNV somatic paired workflow, should I exclude the paired normal sample in PoN?

Previously @shlee said I need to do the germline CNV extraction manually after I get the segment results from both tumor and normal. https://gatkforums.broadinstitute.org/gatk/discussion/24229/call-paired-somatic-cnv#latest

The problem here is when I include my normal sample in the PoN, the final CNV calls for the normal sample seem to have nothing. So I don't have anything to remove from the tumor. I think the program treated most of the points in my normal sample as noise so it removed them, but not all points. I can still see some sparse points on the normal sample plot. It seems to help me remove most of the germline CNVs.

1. Do you recommend to remove the paired normal sample when creating PoN.

2. If I include the paired sample in my PoN, do you think it is good enough to remove germline CNVs?

Segment plot when I include this normal sample in PoN:
normal

The paired tumor sample:
tumor

Best Answers

Answers

  • lzhan140lzhan140 Member

    Hi @shlee,

    could you give me some recommendations?

    Thank you!

  • lzhan140lzhan140 Member

    Hi @slee,

    Yeah I meant you, sorry.

    I used 13 samples for my PoN and 3 of them are replicates, so 10 different individuals. I was running on Terra and I didn't change the eigensamples string (#use all eigensamples in panel by default). The PoN creation was also default (20 eigansamples).

    Would you recommend me to use a small number?

    Thanks.

  • lzhan140lzhan140 Member

    Hi @slee,
    I extracted all the singluar values for my PoN

    252.6883192546395
    241.98341466462037
    240.27440331099712
    239.10785645601126
    236.95645436422564
    233.37090291352203
    228.02019462415257
    222.19674853753116
    218.28387774821473
    215.90399032366327
    212.1380520021574
    206.93633068946113
    165.7532712844405

    It seems it's decreasing all the way. I don't see an "elbow" like you would expect in this:

    So what eigan sample size should I use?

  • lzhan140lzhan140 Member

    Hi @slee,

    Thanks for the recommendations. We just want to make some confident somatic CNV calls. By "confident", it means that we want to be able to tell which ones are somatic and which ones are in the germline. Current PoN, like you said, over-denoised the normal sample, so we are not able to tell which CNVs are germline or somatic.

    I will first try with different PCs.

  • lzhan140lzhan140 Member
    edited July 30

    Hi @slee,

    Here are some results. I tried to denoise one of my normal samples with eigan sample from all, 10, 7 ,4 to 1. Here is what I found: It looks like only when eigan sample = 1 gives me a similar level of sensitivity as the paired tumor. Notice the deletion on chr7. It constantly appears in my paired normal and only when eigan_sample=1 can detect it.

    Do you think I should just use eigan_sample=1 for normals samples and all for tumor samples?

    Another question: I noticed a deletion was called on chr19 no matter the eigan sample = 1, 4 or 10 for the normal sample, but not in the compared tumor sample. However, I do see a small peak at that position in paired tumors. Is it because it is over denoised in tumor (eigan=13 for tumor)? Then, I guess I'd better use eigan=1 for all samples?

    two of paired tumor:

    Thanks!

  • lzhan140lzhan140 Member

    Hi @slee,

    Thanks for your answers. They are super helpful.

    I tried with my noisest tumor sample with eigan= 13 and 0. I found a minium difference, but to increase sensitivity and match with the normal samples, I will still use 0 for all.

    Top: 0; Bottom: 13

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