Hello,
I am new to use cnvkit and i read manual of cnvkit. I did not understand so many things. I want to used cnvkit to calculate copy number in my sequencing data with no normal sample. So i followed the steps that mentioned in manual. In my analysis, i have target file that include all the exon positions. From the target bed file, i got the antitarget file with antitarget command. Can someone explain me that what does mean of antitarget region. As i read, i got that region between target region is is so or not ?? and i also do not understand what is the role of access command ?? Can some one explain me with example ?? i will be grateful for your help.
Thanks
1 answer
Off-target or "antitarget" regions are explained in the CNVkit paper here: http://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1004873
You don't need a matched normal/control sample for the analysis, although it will usually improve your results.
Some regions of the genome are not easily sequenced, e.g. telomeres, centromeres, and some highly repetitive regions. These are masked out with "N" characters in the reference genome FASTA sequence. The remaining regions are called sequencing-accessible regions. CNVkit's access command identifies the locations of the sequencing-accessible regions so that the hard-to-sequence regions are excluded from the analysis.
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First you should understand the algorithm at least briefly. You need to have pairs like tumor vs normal or wild-type vs knock-down vs.. Without pair you can not estimate copy number variations since you calculate the ratio of read counts in log scale across tumor and normal. By doing that, you can estimate the loss and gain in different loci. You can read Varscan paper which is explained good.