Daniel:Notebook/HiResChrPaint/2013-10-23: Difference between revisions

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[[Daniel:Notebook/HiResChrPaint|Back to Calendar]]
[[Daniel:Notebook/HiResChrPaint|Back to Calendar]]


==DpnII Digestion ([[Daniel:Notebook/HiResChrPaint/2013-10-22#DpnII Digestion|Started 10/22/2013]]==
==DpnII Digestion ([[Daniel:Notebook/HiResChrPaint/2013-10-22#DpnII Digestion|Started 10/22/2013]])==


  3. Column Purification, elute with 30 uL EB
  3. Column Purification, elute with 30 uL EB
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'''Gel Image'''
'''Gel Image'''
[[Image:20131023_DpnIIDigest.jpg|600px]]


==Lambda Exonuclease Digestion==
==Lambda Exonuclease Digestion==
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  1. Reaction table
  1. Reaction table


{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext>
|- style="font-size:12pt;font-weight:bold" align="center"
| width="65" height="15" | Sample
| width="65" | BSPP 1AB
| width="65" | BSPP 1CD
| width="65" | BSPP 2AB
| width="65" | BSPP 2CD


|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | ng/uL
| align="center" | 166.5
| align="center" | 168.8
| align="center" | 180.2
| align="center" | 143
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | ug in 25 uL
| align="center" | 4.2
| align="center" | 4.2
| align="center" | 4.5
| align="center" | 3.6
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | uL added
| align="center" | 25
| align="center" | 25
| align="center" | 25
| align="center" | 25
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="30" | uL 10x buffer
| align="center" | 5
| align="center" | 5
| align="center" | 5
| align="center" | 5
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="45" | uL Lambda Exonuclease (5U/uL)
| align="center" | 3
| align="center" | 3
| align="center" | 3
| align="center" | 3
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | uL nfH2O
| align="center" | 17
| align="center" | 17
| align="center" | 17
| align="center" | 17
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | total
| align="center" | 50
| align="center" | 50
| align="center" | 50
| align="center" | 50
|}


  2. Incubate for 1 hour at 37C followed by 10 minute heat inactivation at 75C
  2. Incubate for 1 hour at 37C followed by 10 minute heat inactivation at 75C

Latest revision as of 21:00, 14 November 2013

BSPP Control (Started 10/22/2013)[edit]

Back to Calendar

DpnII Digestion (Started 10/22/2013)[edit]

3. Column Purification, elute with 30 uL EB
4. Measure concentrations in nanodrop

Nanodrop Results[edit]

Sample BSPP 1AB BSPP 1CD BSPP 2AB BSPP 2CD
ng/uL 166.5 168.8 180.2 143
ug in 30 uL 5.0 5.1 5.4 4.3
total   10.1   9.7

TBE Gel[edit]

1. Lanes; (5.5 uL TBE; 2 uL 6x buffer, 0.5 uL sample; load 7.2 uL for all)


Lane 1 2 3 4 5 6 7 8 9 10 11 12
Sample 10bp ladder BSPP 1AB BSPP 1CD BSPP 2AB BSPP 2CD
2. Run for 23 minutes at 220V
3. Stain with 3 uL SYBR gold for 2 minutes

Gel Image

File:20131023 DpnIIDigest.jpg

Lambda Exonuclease Digestion[edit]

1. Reaction table
Sample BSPP 1AB BSPP 1CD BSPP 2AB BSPP 2CD
ng/uL 166.5 168.8 180.2 143
ug in 25 uL 4.2 4.2 4.5 3.6
uL added 25 25 25 25
uL 10x buffer 5 5 5 5
uL Lambda Exonuclease (5U/uL) 3 3 3 3
uL nfH2O 17 17 17 17
total 50 50 50 50
2. Incubate for 1 hour at 37C followed by 10 minute heat inactivation at 75C
3. Pool samples
4. ssDNA column purification, elute with 20 uL nfH2O
5. Measure Concentration in Nanodrop

ssNanodrop Results[edit]