AlanFung:LabNotes/Capturing/2012-8-10: Difference between revisions

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>Alan6017518
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===Prepare Exo I and III Mix===
===Prepare Exo I and III Mix===
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8<\hiddentext>
|- style="background-color:#538DD5;font-size:8pt" align="center"
|style="font-weight:bold;text-decoration:underline" width="121" height="13" | Exo I & III for 20ul Rxn
|style="font-weight:bold;text-decoration:none" width="39" | 1X
| width="33" | 16.5x
|- style="font-size:8pt" align="center"
| height="13" | Exo I (20U/ul)
| align="center" | 1
| align="center" | 16.5
|- style="font-size:8pt" align="center"
| height="13" | ExoIII (200U/ul)
| align="center" | 1
| align="center" | 16.5
|- style="font-size:8pt" align="center"
| align="center" height="13" | &nbsp;
| align="center" | &nbsp;
| align="center" | &nbsp;
|- style="background-color:#538DD5;font-size:8pt" align="center"
|style="font-weight:bold;text-decoration:underline" height="13" | Exo I & III for 50ul Rxn
|style="font-weight:bold;text-decoration:none" | 1X
| 19.8x
|- style="font-size:8pt" align="center"
| height="13" | Exo I (20U/ul)
| align="center" | 2.5
| align="center" | 49.5
|- style="font-size:8pt" align="center"
| height="13" | ExoIII (200U/ul)
| align="center" | 2.5
| align="center" | 49.5
|}

Revision as of 00:33, 11 August 2012

Capturing Remaining Gimelbrant lab Sample

  • Add 2 drops of mineral oil to prevent evaporation
ChIPK36me3 Concentration ng/ul Vol    Vol of probes 10X Ampligase Buffer H2O Total
GM12878 0.90 40.00 0.45 2 7.6 50
P10 1.20 40.00 0.60 2 7.4 50
DF2 0.70 40.00 0.35 2 7.7 50
H0 1.00 40.00 0.50 2 7.5 50
H7 0.60 40.00 0.30 2 7.7 50
H14 0.50 40.00 0.25 2 7.8 50
H16 0.60 40.00 0.30 2 7.7 50
                
ChIPK27me3 Concentration ng/ul Vol    Vol of probes 10X Ampligase Buffer H2O Total
GM12878 1.20 40.00 0.60 2 7.4 50
GM12878 Round 2 2.86 35.00 1.25 2 11.8 50
P10 1.30 40.00 0.65 2 7.4 50
DF2 2 40.00 1.00 2 7.0 50
H0 2.4 40.00 1.20 2 6.8 50
H7 1.2 40.00 0.60 2 7.4 50
H14 0.5 40.00 0.25 2 7.8 50
H16 1.3 40.00 0.65 2 7.4 50
                
WCE Concentration ng/ul Vol    Vol of probes 10X Ampligase Buffer H2O Total
GM12878 35.39 5.65 2.49 2 9.9 20
GM12878 Round 2 2.86 35.00 1.25 2 11.8 50
P10 35.39 5.65 2.49 2 9.9 20
P10 Round 2 2.86 35.00 1.25 2 11.8 50
DF2 35.39 5.65 2.49 2 9.9 20
DF2 Round 2 2.86 35.00 1.25 2 11.8 50
H0 35.39 5.65 2.49 2 9.9 20
H7 35.39 5.65 2.49 2 9.9 20
H14 35.39 5.65 2.49 2 9.9 20
H16 35.39 5.65 2.49 2 9.9 20
                
cDNA Concentration ng/ul Vol    Vol of probes 10X Ampligase Buffer H2O Total
GM 38.12 5.25 2.49 2 10.3 20
H0 38.12 5.25 2.49 2 10.3 20
H7 38.12 5.25 2.49 2 10.3 20
H14 38.12 5.25 2.49 2 10.3 20
H16 38.12 5.25 2.49 2 10.3 20
                
gDNA Concentration ng/ul Vol    Vol of probes 10X Ampligase Buffer H2O Total
H7 38.12 5.25 2.49 2 10.3 20
H14 38.12 5.25 2.49 2 10.3 20
H16 38.12 5.25 2.49 2 10.3 20
  • Make sure we have enough reagent before start,
  • need 50ul probes, check
  • need 66ul 10X Ampligase Buffer,

Prepare SLN Mix

  • Dilute 1mM dNTP 1 in 100 to 10uM dNTP
SLN Mix for 20ul Rxn 1x 16.5x
dNTP (10uM) 0.5 8.25
Stoffel (10U/ul) 0.2 3.30
Ampligase (5U/ul) 0.2 3.30
H2O 0.1 1.65
   
SLN Mix for 50ul Rxn 1x 19.8x
dNTP (10uM) 1.25 24.75
Stoffel (10U/ul) 0.5 9.90
Ampligase (5U/ul) 0.5 9.90
H2O 0.25 4.95

Prepare Exo I and III Mix

Exo I & III for 20ul Rxn 1X 16.5x
Exo I (20U/ul) 1 16.5
ExoIII (200U/ul) 1 16.5
     
Exo I & III for 50ul Rxn 1X 19.8x
Exo I (20U/ul) 2.5 49.5
ExoIII (200U/ul) 2.5 49.5