Comments on: PCR – Oly RAD-seq Prep Scale PCR http://onsnetwork.org/kubu4/2015/10/13/pcr-oly-rad-seq-prep-scale-pcr/ University of Washington - Fishery Sciences - Roberts Lab Tue, 09 Apr 2019 18:37:58 +0000 hourly 1 http://wordpress.org/?v=4.0 By: Restriction Digest – Oly gDNA for RAD-seq w/AlfI | Sam's Notebook http://onsnetwork.org/kubu4/2015/10/13/pcr-oly-rad-seq-prep-scale-pcr/#comment-2350 Thu, 29 Oct 2015 00:11:16 +0000 http://onsnetwork.org/kubu4/?p=1730#comment-2350 […] The previous attempt at making these RAD libraries failed during the prep-scale PCR, likely due to a discrepancy in the version of the Meyer Lab protocol I was following, so I have to start at the beginning to try to make these libraries again. […]

]]> By: Troubleshooting – Oly RAD-seq | Sam's Notebook http://onsnetwork.org/kubu4/2015/10/13/pcr-oly-rad-seq-prep-scale-pcr/#comment-2257 Fri, 23 Oct 2015 22:50:30 +0000 http://onsnetwork.org/kubu4/?p=1730#comment-2257 […] After the failure of the prep-scale PCR for the RAD library construction, Katherin Silliman pointed out a potential problem (too much dNTPs). This was odd because I was following the Meyer Protocol and I used what was indicated. […]

]]> By: kubu4 http://onsnetwork.org/kubu4/2015/10/13/pcr-oly-rad-seq-prep-scale-pcr/#comment-2180 Sat, 17 Oct 2015 02:54:38 +0000 http://onsnetwork.org/kubu4/?p=1730#comment-2180

Hmmm, the Meyer Lab protocol says 20uL of 10mM dNTPs (which is what I used). Where are you getting your numbers from? Thanks for looking this over, btw.

]]> By: Katherine Silliman http://onsnetwork.org/kubu4/2015/10/13/pcr-oly-rad-seq-prep-scale-pcr/#comment-2169 Fri, 16 Oct 2015 17:08:36 +0000 http://onsnetwork.org/kubu4/?p=1730#comment-2169 Did you use 10mM DNTPs or 1mM? For the prepscale PCR it should either be 20 uL of 1mM or 2 uL of 10mM per sample.

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