Daniel:Notebook/HiResChrPaint/2013-11-14: Difference between revisions

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>Djacobse
(Created page with "=Positive Control 2= I'm going to start my work on positive control 2 today, amplifying all 4 probe sets and eventually coupling them to a single dye (Alexa 488). Since I've...")
 
>Djacobse
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===qPCR Results===
===qPCR Results===
[[Image:2013-11-14-V6S1-V6S2.png|500px]]


===Nanodrop Results===
===Nanodrop Results===

Revision as of 19:47, 14 November 2013

Positive Control 2

I'm going to start my work on positive control 2 today, amplifying all 4 probe sets and eventually coupling them to a single dye (Alexa 488). Since I've found I work best only amplifying 2 sets at a time, I'll amplify V6S1 and V6S2 first, and amplify S3 and S4 starting tomorrow or Saturday.

qPCR Amplification

1. Reaction table
  10 nM OligoPool 2X Kapa SYBR qPCR MM 100 uM AP1V61U-Amino 100 uM AP2V6 H20 Total Volume
per rxn 0.4 50 0.4 0.4 48.8 100
V6S1 (12.5x) 5 625 5 5 610 1250
V6S2 (12.5x) 5 625 5 5 610 1250


2. Aliquot 100 uL into each of 12 qPCR tubes 
3. PCR protocol:
    i. PCR 3 min 95C
    ii. 95C 3 sec
    iii. 55C 30 sec 
    iv. 72C 20 sec
    v. plate read
    vi. goto ii x17
    vii. 72C 2 min
    viii. 16C hold
4. Pooled 3 samples together each (300 uL each, 4 pools per set, 8 total)
5. Column Purification, elute with 40 uL EB
6. Measure concentrations in nanodrop


qPCR Results

File:2013-11-14-V6S1-V6S2.png

Nanodrop Results