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Hi,
I wanted to ask how to calculate the expected depth coverage in 454 if I know the size of the sequenced genome?
I wanted to ask how to calculate the expected depth coverage in 454 if I know the size of the sequenced genome?
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Your local sequencing center should be able to tell you how much raw sequence data a plate (or half a plate, 1/4, 1/8 or even 1/16 of a plate) would give you (in base pairs), so divide this by the estimated genome size. That will will give you an upper bound on the coverage assuming uniformity of coverage, linear amplification, no contamination, etc.
If you want more practical comments, I suggest you tell the forum what kind of organism it is and the estimated genome size. The GC% might also be important... -
GC content
Why is GC content important? how to consider it?Last edited by dina; 10-03-2009, 06:27 AM.Comment
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Well, not directly, but extreme GC values could cause trouble at several steps.Comment
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GC
So, what would you sugggest? I have to check the GC content of the whole genome before the sequencing? and if it is high , what can I so?Comment
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If you have any genes or ESTs already sequenced you should be able to guess the GC%, or even look at related organisms. If it is extreme then some background reading of the literature might be in order - but you'll probably be fine.Comment
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Love this - good data definitely starts from good input, and poor input can only give relatively poor data. I particularly like the mention of Nanodrop/absorbance based methods for quantification. It's such a toss up if you'll get an accurate reading or what amounts to a randomly generated number, and a lot of library/sequencing related issues can be traced back to poor quant.07-01-2026, 11:43 AM
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I’m not a sequencing expert. I’m a purification scientist who uses NGS to evaluate workflows my group develops. With this perspective, we think about the sample first and the NGS workflow second. The sequencer is an exceptionally honest reporter, but it can only report on what you give it, so whether you get clean, interpretable data from an NGS workflow is largely determined before you begin.
Here are nine questions we think about, in roughly the order they matter, before...
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