Calendar/2012-7-13: Difference between revisions

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{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8<\hiddentext>
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8<\hiddentext>
|- style="background-color:#B7DEE8;font-size:8pt;font-weight:bold" align="center"
|- style="background-color:#B7DEE8;font-size:8pt;font-weight:bold" align="center"
|style="float: right;"
| style="float: right;"
|style="text-decoration:underline" width="115" height="15" | Content
|style="text-decoration:underline" width="115" height="15" | Content
| width="110" | Volume Per Reaction
| width="110" | Volume Per Reaction

Revision as of 01:32, 16 July 2012

Library Construction of Dr. Yi Zhang's DiP libraries Sample

Samples Preparation Protocol from Li

File:0712 DIP-seq samples.pdf

  • Received samples from Li on Jul 13 2012, 12 samples total in a yellow box, transfer to -20C

qPCR with N2 Barcodes

DIP Sample N2 Barcode
shCONT-input 1
shCONT-IgG 2
shCONT-5mC 3
shCONT-5hmC 4
shCONT-5fc 5
shCONT-5caC 6
shTDG-input 7
shTDG-IgG 8
shTDG-5mC 9
shTDG-5hmC 10
shTDG-5fC 11
shTDG-caC 12
style="float: right;" Content Volume Per Reaction MM
Template 10 Do not add to MM
5uM PCR F+R Barcode 1 Do not add to MM
50X SYBG 0.2 2.64
2X PhusionHF MM 25 330
H20 13.8 182.16
Total Volume 50 660
  • Add 20ul 10uM N2-bar-code primers into strip tube
  • Add 20ul 10uM PCR_F primers and mix
  • We have 5uM F+R N2 Ind primer (Store at 4C Alice yellow 0.2ul tube rack)
  • Add 10ul template (for low volume sample add 5ul sample and add 5ul h2o) and 1ul barcode primers (using multi channel pipette) to strip tube
  • Add 39ul MM to each well
  98C 30sec 
  (98C 10sec -> 63C 20sec-> 72C 30sec) x 20 cycles
  72C 2min  
  15C hold.

qPCR Curves

Gel quant

Qubit

DIP Sample N2 Barcode Sample Concentration ng/ul Volume ul Amount ng
shCONT-input 1 3.62 30 108.6
shCONT-IgG 2 2.36 30 70.8
shCONT-5mC 3 2.64 30 79.2
shCONT-5hmC 4 3.54 30 106.2
shCONT-5fc 5 3.58 30 107.4
shCONT-5caC 6 3.82 30 114.6
shTDG-input 7 3.54 30 106.2
shTDG-IgG 8 2.4 30 72
shTDG-5mC 9 3.06 30 91.8
shTDG-5hmC 10 4.04 30 121.2
shTDG-5fC 11 3.9 30 117
shTDG-caC 12 3.18 30 95.4