Revision as of 21:45, 6 June 2013 by >Djacobse
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Summary of DpnII Digestion Reactions
Alpha Group (aa-dUTP present)
Sample
|
V4S2A
|
V4S2B
|
V4S2C
|
V4S2D
|
V6S2A
|
V6S2B
|
V6S2C
|
V6S2D
|
ng/uL
|
121.5
|
127.1
|
119.1
|
121.8
|
136.7
|
122.4
|
119.2
|
147.7
|
ug left
|
3.6
|
3.8
|
3.6
|
3.7
|
4.1
|
3.7
|
3.6
|
4.4
|
total
|
|
|
|
14.7
|
|
|
|
15.8
|
Beta Group (no aa-dUTP)
Sample
|
V4S2A
|
V4S2B
|
V4S2C
|
V4S2D
|
V6S2A
|
V6S2B
|
V6S2C
|
V6S2D
|
ng/uL
|
126.4
|
122.6
|
108.5
|
112.2
|
129.7
|
112.5
|
140.4
|
128.7
|
ug left
|
3.8
|
3.7
|
3.3
|
3.4
|
3.9
|
3.4
|
4.2
|
3.9
|
total
|
|
|
|
14.1
|
|
|
|
15.3
|
Lambda Exonuclease Digestion (Alpha Group)
1. Prepare samples
Sample
|
V4S2A
|
V4S2B
|
V4S2C
|
V4S2D
|
V6S2A
|
V6S2B
|
V6S2C
|
V6S2D
|
ng/uL sample
|
121.5
|
127.1
|
119.1
|
121.8
|
136.7
|
122.4
|
119.2
|
147.7
|
uL sample
|
25
|
25
|
25
|
25
|
25
|
25
|
25
|
25
|
ug sample
|
3.0375
|
3.1775
|
2.9775
|
3.045
|
3.4175
|
3.06
|
2.98
|
3.6925
|
uL 10x buffer
|
5
|
5
|
5
|
5
|
5
|
5
|
5
|
5
|
uL lambda exonuclease (5U/uL)
|
3
|
3
|
3
|
3
|
3
|
3
|
3
|
3
|
uL nf H20
|
17
|
17
|
17
|
17
|
17
|
17
|
17
|
17
|
total
|
50
|
50
|
50
|
50
|
50
|
50
|
50
|
50
|
2. Incubate at 37C for 1 hour followed by 10 minute heat shock at 75C
3. Pool samples into single tube (4x50uL->1x200uL)
4. ssDNA column purification