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Forum: Illumina/Solexa 03-23-2011, 08:52 AM
Replies: 32
Views: 24,173
Posted By JUdw
End-repair, A-tailing and adpter ligation is...

End-repair, A-tailing and adpter ligation is combined in the Nextera sample prep protocol:

http://www.epibio.com/item.asp?ID=566
Forum: Illumina/Solexa 03-23-2011, 02:08 AM
Replies: 32
Views: 24,173
Posted By JUdw
I don't think you can use the standard Ampure...

I don't think you can use the standard Ampure cleaning protocol, it removed most of the <75 bp fragments when I used it. 70bp DNA in combination with ~75bp adapters might work however, see attachment.
Forum: Illumina/Solexa 08-19-2010, 08:57 AM
Replies: 43
Views: 135,976
Posted By JUdw
The salmon sperm binds aspecific to the baits and...

The salmon sperm binds aspecific to the baits and gets replaced by target DNA during hybridisation? Then you get lots of salmon sperm in your enriched fraction, since there is an excess of baits. Or...
Forum: Illumina/Solexa 08-19-2010, 02:48 AM
Replies: 43
Views: 135,976
Posted By JUdw
In the article 'Enrichment of sequencing targets...

In the article 'Enrichment of sequencing targets from the human genome by solution hybridization', Tewhey et al. 2009. They use the Agilent SureSelect baits and say in m&m:

"500 ng of each genomic...
Forum: Illumina/Solexa 07-27-2010, 05:23 AM
Replies: 19
Views: 10,270
Posted By JUdw
1ul [25uM] in 50ul PCR reaction, [final] = 0.5uM

1ul [25uM] in 50ul PCR reaction, [final] = 0.5uM
Forum: Sample Prep / Library Generation 07-27-2010, 01:12 AM
Replies: 10
Views: 5,167
Posted By JUdw
Did you tile your bait design? In another...

Did you tile your bait design?

In another thread I posted a question on the bait to sample ratio (http://seqanswers.com/forums/showthread.php?t=6091)

From your answer I get that this ratio is...
Forum: Sample Prep / Library Generation 07-26-2010, 06:15 AM
Replies: 5
Views: 7,210
Posted By JUdw
I already have about 500ng of template, so i...

I already have about 500ng of template, so i think I don't really need to do the pre-hyb PCR. For qPCR the pre-hyb PCR is however required, since these primers only anneal to the extended PCR...
Forum: Sample Prep / Library Generation 07-23-2010, 06:20 AM
Replies: 1
Views: 3,925
Posted By JUdw
In-solution SureSelect capture library to prepped sample ratio

Im wondering about the in-solution SureSelect library to prepped sample ratio. The SureSelect protocol states The ratio of SureSelect capture library to prepped library is critical for successful...
Forum: Sample Prep / Library Generation 05-14-2010, 07:55 AM
Replies: 5
Views: 7,210
Posted By JUdw
Could you further reduce the pre-hyb PCR cycle...

Could you further reduce the pre-hyb PCR cycle number, e.g to 3 rounds of PCR using PE primers?

I suppose theoretically 3 rounds is the minimum, since you need the symmetric PCR product mentioned...
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