Noi/NOTES/2014-10-22

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Revision as of 16:19, 22 October 2014 by >Noi (Created page with "= Preparation of WGBS of cfDNA from plasma using KAPA Hyper Prep kit = * Rational: Will be added '''Sample list'''<br> '''No.''' '''Sample IDs''' '''Conc. (ng/ul)''' #1 NC-2...")
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Preparation of WGBS of cfDNA from plasma using KAPA Hyper Prep kit

  • Rational: Will be added

Sample list

No.	Sample IDs	Conc. (ng/ul)
#1	NC-2	        0.325
#2	NC-3	        0.538
#3	6P-2	        1.10
#4	6P-3	        1.57
  • Note: I will reduce the reaction volume to be half of the instruction and reduce the reagent volume/amount proportionally.
  • Actual reaction set up:
- 50ul of fragmented DNA
- 7ul of End repair & dA-tailing buffer
- 3ul of End repair & dA-tailing enzyme mix
- Total reaction: 60ul

End repair & dA-tailing

End repair & dA-tailing reaction mix

Components Volume half rxn 5.2x rxn
End repair/dA-tailing buffer 7.00 3.50 18.20
End repair/dA-tailing enzyme mix 3.00 1.50 7.80
Total 10.00 5.00

Reaction set up

Sample IDs Conc. (ng/ul) Volume for 1ng (ul) End repair/dA-tailing rxn mix H2O (ul) Total (ul)
NC-2 0.33 3.08 5.00 21.92 30.00
NC-3 0.54 1.86 5.00 23.14 30.00
6P-2 1.10 0.91 5.00 24.09 30.00
6P-3 1.57 0.64 5.00 24.36 30.00
NTC 0.00 0.00 5.00 25.00 30.00
- Mix the reaction mix well
- 20C, 30min
- 65C, 30min,
- Hold 4C
- Proceed to Adaptor ligation immediately

Adaptor ligation