Daniel:Notebook/HiResChrPaint/2013-6-3

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Probe Prep 3 (Started 5/29/2013)[edit]

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Gel for DpnII Digestion[edit]

Looking to see if beta group (no aa-dUTP) has been digestion efficiency than alpha group (with aa-dUTP).
1. Set up 0.5 uL sample, 2 uL 6x dye, 5.5 uL TBE
Lane 2 3 4 5 6 7 8 9 10 11
Sample 10 bp ladder V4S2 V6S2 V4S2* V6S2* 10 bp ladder V4S2 V6S2 V4S2* V6S2*
  • denotes DNA without aa-dUTP (beta group)
2. Run for 25 minutes at 220V
3. Stain for 3 minutes with 3 uL SYBR gold

Gel Image[edit]

File:DpnDigest 060313 aadUTPvsno.jpg

Discussion[edit]

From this image it is not clear whether or not the aa-dUTP is making a difference.  I'll try again with the 16 hour digestion to see if it is any
better.

DpnII Digestion Test (16 hours)[edit]

Only running 1/4 of DNA I have for each sample to compare DpnII reaction yield using aadUTP DNA and normal DNA.  Using longer incubation time.
1. Set up 
Sample uL sample uL Dpn II (50 U/uL) uL 10x buffer uL H20 total
alpha 4-B 35 2 5 8 50
alpha 6-B 35 2 5 8 50
beta 4-B 35 2 5 8 50
beta 6-B 35 2 5 8 50
2. Incubate in thermocycler at 37C for 16 hours; heat shock 20 minutes at 65 C
3. Continued 6/4/2013