Hii, Greeting Everyone
Currently i am working on Next Generation Sequencing, will anyone suggest me the depth/coverage needed for a sequence obtained through any platform of NGS particularly Illumina Miseq.
It depends upon the type of sequencing you are doing.
For detecting human genome mutations, SNPs, and rearrangements, publications often recommend from 10× to 30× depth of coverage, depending on the application and statistical model.
For RNA sequencing, researchers usually think in terms of numbers of millions of reads to be sampled. Detecting rarely expressed genes often requires an increase in the depth of coverage.
For ChIP-Seq (chromatin immunoprecipitation sequencing), publications often recommend coverage of around 100x.
Thank you Mr. Abdul Rafay Khan, for your valuable suggestion, actually i am targeting hyper variable region of 16S rRNA gene (DNA targeting sequence ), so in this case what will be the recommended depth.
Hi Rajeev,
in addition to the good advices from Abdul, I would add that the depth/coverage depends also on your samples, the quality of them (FFPE samples need higher coverage), depending if you're searching for somatic or constitutional variants, if you're searching to follow clones and so on...
targeting sequencing will allow you easily to reach a 100 to 1000x coverage, which is a good base to work on.
fred
Thank you frederic but as I already mentioned I am targeting hypervariable region of 16S rRNA gene, so what depth may provide good quality data, since if depth is high the assembly of sequences will be easy.
Hi Rajeev,
if you have data on the variability and percents of clones it should help you to decide. In a project I've been implicated we decided to go to a 100.000x coverage to follow the clones and decide further the coverage we would fix for the following patients.
fred
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