Dinh/NOTES/2010-7-3: Difference between revisions

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>Dinh
m (Created page with '==Sanger sequencing of PCR product with probe linker== ===Primers=== LinkerR-LMNA-F: GTT GGA GGC TCA TCG TTC CTA TTT TGG GCC TGA GTG GTC AGT C LinkerL-LMNA-R: GAC CCG CCT GCA …')
 
>Dinh
mNo edit summary
Line 20: Line 20:
  94C  2min
  94C  2min
  94C  30s
  94C  30s
  57C   40s
  62C   40s
  72C  40s
  72C  40s
  Go to 2 for 7x
  Go to 2 for 7x

Revision as of 20:58, 2 July 2010

Sanger sequencing of PCR product with probe linker

Primers

LinkerR-LMNA-F: GTT GGA GGC TCA TCG TTC CTA TTT TGG GCC TGA GTG GTC AGT C
LinkerL-LMNA-R: GAC CCG CCT GCA GGA TTT GGC AGG CAG ATG TTA TCG AGG TCC GAC
Product length: 373+32bp = 405bp
Position: chr1:156,108,195-156,108,590

PCR

Template (1-2ng/ul): 3ul
F primer (10uM)    : 1ul
R primer (10uM)    : 1ul
H20                : 20ul
Taq 2x MM          : 25ul
-----------------------
                   50ul
Templates used: BJ-ips, HGPS-ips, HGPS72
Program: 
94C   2min
94C   30s
62C   40s
72C   40s
Go to 2 for 7x
94C   30s
72C   40s
Go to 7 for 23x
72C   3min
15C   forever
Purify by 0.9x volumes of Agencourt beads.
  • Quantification by Nanodrop


  • Premix for Genewiz sequencing:

Results