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Old 05-06-2010, 10:34 PM   #1
mitchelS
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Default Gel-size selection in NimbleGen exon capture

Hey,

just wondering if anyone has tried omiting the gel-size selection following Illumina adaptor ligation prior to commemcing the LM-PCR step in the NimbleGen in-solution Exon capture protocol?

If so does the residual adaptor dimers have any marked effect on the downstream data?

Cheers,

Mitch.
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Old 06-27-2012, 07:36 AM   #2
Liting
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Hi,mitchels
I am tring the gel-size selection following Illumina adaptor ligation prior to commemcing the LM-PCR step in the NimbleGen in-solution Exon capture protocol。But we can't get enough PCR products after LM-PCR(8 cycles)。Are you successfully with the protocal?
Thank you!
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Old 10-24-2012, 11:15 AM   #3
apeterson
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I have found that the skipping the gel size selection can result in increased primer dimers in the pre-hybridization PCR. The Illumina protocols suggest an extra magnetic bead clean after ligation, this seems to help with the PCR.
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Old 10-26-2012, 09:50 AM   #4
csquared
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Size selection is definitely not needed by gel, but a bead-based size selection to get rid of adaptor dimer is really helpful. A single 1x bead purification will get rid of stuff below 200nt prior to library amplification.
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