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  • exome sequencing coverage

    Hello
    I am designing the exome sequencing of breast cancer tissues.
    The tissues would be FFPE (Formalin Fixed Paraffin Embedded).
    I wonder what level of coverage would be good for rare SNP or structural variations in exome sequencing.
    I was thinking 40X, however, I found a few papers using 100X~200X exome sequencing.
    Plus, any specific caution for FFPE tissues which are likely to include degraded DNA ?
    Please help me !

    Thank you

  • #2
    Hi,

    I guess you will find more information on coverage in other sections of the forum. But anyway, this is an example of how coverage can look like:

    TARGET COVERAGE (EXOME)
    >0 ≥8x ≥20x ≥30x ≥100x
    99,44% 93,51% 81,59% 68,3% 11,7%

    The "exome" is not really the complete exome as the targeted capture strategies used dont capture the COMPLETE exome. You will see that some genes are hardly covered at all, due to this.

    A rare SNP will need better coverage/reads to be certain, but I would anyhow use traditional Sanger sequencing to verify a rare SNP after exome-sequencing. 40x sounds good, even 35x could do it. But be careful not to throw away exciting SNPs because of the quality, check with Sanger!

    For the other part, FFPE, I am sorry that I cant help you. Hope this helps in some way.

    C

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