[NextGenSeq]

Quick summary of Illumina exome data
We get ~120 million reads per lane of HiSeq
The mean exome coverage was about 150X. Thus you can multiplex 2 to 3 whole exomes per lane.
We found our mutation so it worked pretty well.

[Geneus]

Quick summary of Illumina exome data
We get ~120 million reads per lane of HiSeq
The mean exome coverage was about 150X. Thus you can multiplex 2 to 3 whole exomes per lane.
We found our mutation so it worked pretty well.

Thank you...very useful information. Presumably, since each kit allows up to 6 indexes one could choose 50X coverage and do 6 exomes per lane, correct?

[DMO]

Cluster Density

When you get 120million reads/lane, what does your cluster density look like?

Quick summary of Illumina exome data
We get ~120 million reads per lane of HiSeq
The mean exome coverage was about 150X. Thus you can multiplex 2 to 3 whole exomes per lane.
We found our mutation so it worked pretty well.

[asiniard]

Thank you...very useful information. Presumably, since each kit allows up to 6 indexes one could choose 50X coverage and do 6 exomes per lane, correct?

Has anyone tried this?

[GW_OK]

With the Truseq system we do 6 exomes per pre-capture pool, 3 lanes per pool on the Hiseq and get about 50x coverage each.

We're happy with the data so far.

[asiniard]

We are runnning several Illumina whole exome lanes now on the HiSeq. Once I analyze the data I'll post the coverage here.

One thing I don't like is that the protocol is very long and tedious. You do two rounds of amplification. Also, you first use the TruSeq DNA kit to make the initial library before the whole exome capture. In our hands about half the libraries didn't yield enough to continue on with the whole exome capture (you need a minimum of 500 ng).

What do you use to quantitate your libraries?

[NextGenSeq]

We use NanoDrop and KAPA QPCR

[dottomarco]

I guess it is with the V2 reagents right NextGenSeq?? TrueSeq V3 should produce 375 millions paired end reads oer lane.. So now it should be possible to get 6 exomes per lane with a pretty decent coverage (75X).

Did someone try or is trying the new TrueSeq sample prep protocol that allows to skip the gel size selection? I think it was released a couple of weeks ago (regents are the same as before)

Quick summary of Illumina exome data
We get ~120 million reads per lane of HiSeq
The mean exome coverage was about 150X. Thus you can multiplex 2 to 3 whole exomes per lane.
We found our mutation so it worked pretty well.

[ashchin]

Hi All,

What amount of exome do you get after 12 cycles of PCR, after hybridization using the agilent human all exon version 2.0. I have input 500ng of illumina library for capture hybridization and ended up getting around 700 ng (Based on the readings from Bioanalyzer)

PS : I have taken 28 ul of captured DNA into PCR.

Thank you,
Ashwini