Exome sequencing? WHY and WHY not?

[kushald]

Unlike what was predicted by many that WGS of human genome will take over exome sequencing, 100s of 1000s of exomes are being sequenced around the world. In simple terms the X10 and X5 (terminated after a prenatal test? not sure) seems to be a secret community out of mainstream.

Following are the most used applications of exome sequencing, please add more, Thank you
PS: also please add if any other kind of analysis can be performed on exome data.

Genetic studies - families for Mendelian inheritance
Population studies
Cancer (low frequency and high frequency mutations)
Consumer Genomics / Personal genomics
Look forward to your comments, Thanks

[NextGenSeq]

A genome sequence will be 1K or less by the end of this year. It will be cheaper to sequence the whole genome.
The MinION mark1 will supposedly generate 40 GB per day this summer. That's over 30X coverage of the genome.

[Brian Bushnell]

A genome sequence will be 1K or less by the end of this year. It will be cheaper to sequence the whole genome.
The MinION mark1 will supposedly generate 40 GB per day this summer. That's over 30X coverage of the genome.

40Gbp/3Gbp = 13X. And as for a MinION producing quality data at that rate, I will believe it when I see it.

[Ola]

With all channels occupied at all times it would give ~22 gb/day for 1d reads. Run time can be up to three days which would make it closer to 30x. As for quality, it could increase with higher speed, but real-world usable throughput is likely going to be much less than the maximum, based on current results something like 15-20 gb total looks more reasonable. PromethIon and MinIon mk2 will probably take us well below $1000 but there may still be some use for exome seq for low freq variants even then.

[jwfoley]

I didn't think Oxford Nanopore was even pretending that the MinION would be a realistic platform when all you care about is coverage. Last I heard, they were saying $1000 per gigabase; HiSeq is already well below $50, and sequences hundreds of Gb a day. Not to mention Nanopore reads will probably be longer than many exons, meaning it'll be hard to get the coverage where you want it.

MinION etc. is a niche platform for long reads and fast turnaround, like Pac Bio, or for field work, unlike Pac Bio.

[kushald]

When is the MinION out in the market? Any idea on the pricing?

[Ola]

When is the MinION out in the market? Any idea on the pricing?

It is on the market but you have to apply to join MAP. What they have said about pricing is an initial $270 per flowcell (3h run time at 500 bases/s).

[Ola]

I didn't think Oxford Nanopore was even pretending that the MinION would be a realistic platform when all you care about is coverage. Last I heard, they were saying $1000 per gigabase; HiSeq is already well below $50, and sequences hundreds of Gb a day. Not to mention Nanopore reads will probably be longer than many exons, meaning it'll be hard to get the coverage where you want it.

MinION etc. is a niche platform for long reads and fast turnaround, like Pac Bio, or for field work, unlike Pac Bio.

It will not sequence reads longer than the input fragments, if you give it short fragments you will just get more reads (unlike PacBio) so there is no reason you could not do exome capture if you wanted to. Not cost-effective compared to illumina yet (unless you factor in the cost of the instrument).

[kushald]

Does any one have an idea on the Agilent V5 kit? Does it cover the non-coding regions? If not, which other kits cover the non-coding region for Exome study?

[NextGenSeq]

I didn't think Oxford Nanopore was even pretending that the MinION would be a realistic platform when all you care about is coverage. Last I heard, they were saying $1000 per gigabase; HiSeq is already well below $50, and sequences hundreds of Gb a day. Not to mention Nanopore reads will probably be longer than many exons, meaning it'll be hard to get the coverage where you want it.

MinION etc. is a niche platform for long reads and fast turnaround, like Pac Bio, or for field work, unlike Pac Bio.

From Clive Brown's blog

Just thinking out loud here...



Fast mode might well get you 40-90Gbases on one flowcell. This would be a best case and fully tuned up, possibly we would extend the lifetime of the flowcell by 24 hrs.

[Shiwali Goyal]

Im doing whole exome sequencing on Ion torrent platform. I started with 1.4microgram/microliter of stock DNA. The fragmentation of the DNA was done correctly as I can see the smear on E-gel near 130-150basepair. Next I proceed with the adapter ligation of the library and then to the size selection step of the library preparation. On library preparation step I picked up the library of 210 basepair size. Then I proceed with the size selected library amplification. When I checked the Qubit concentration of my amplified library, it was below 4 ng/microliter. This concentration must be near 10 or 11 ng/microlitre. Can anyone tell me where the flaw is in this whole experiment?