Jie:LabNotes/ASE/2008-9-22: Difference between revisions
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98C 30S -> (98C 10S -> 58C 20S -> 72C 20S) x 8 -> (98C 10S -> 72C 20S) x 8 -> 72C 5 min -> 15C hold. | 98C 30S -> (98C 10S -> 58C 20S -> 72C 20S) x 8 -> (98C 10S -> 72C 20S) x 8 -> 72C 5 min -> 15C hold. | ||
[[Image:20080924_Yuan Gao sample SNP84k capture.jpg]]20080924_Yuan Gao sample SNP84k capture | |||
Qiaquick column purification. Yields: | |||
G2: 26.1ng/ul (132nM) x 30ul | |||
2B: 26.8ng/ul (140nM) x 30ul | |||
3B: 40.7ng/ul (213nM) x 30ul | |||
H1: 33.4ng/ul (175nM) x 30ul | |||
H2: 43.3ng/ul (227nM) x 30ul | |||
Sent 15ul products to Yuan Gao on 09/24/08. |
Latest revision as of 21:32, 24 September 2008
padlock capture of sample from Yu Gao with eSNP84K[edit]
capture with the padlock probe made on 08-03-27.
tube 1: G2 (~0.25ug/ul) tube 2: 2B (~3.5ng/ul) tube 3: 3B (~3.5ng/ul) tube 4: H1 (~5ng/ul) tube 5: H2 (~100ng/ul) tube 6: Jurkat DNA (5ng/ul) tube 7: negative control reaction system tube 1 x7 H2O 6.3ul 44.1ul 10x Ampligase Buffer 1ul 7ul probe(2008-03-27,30ng/ul) 3ul 21ul template 2ul 2ul each
ddH2O Ligase Buffer Ligase Amplitaq dNTP 2U/ul AmpliTaq Stoffel fragment; 0.5U/ul AmpLigase; 0.8nM dNTP 0.2ul 0.1ul 0.1ul 0.2ul (2nM)0.4ul x20 4ul 2ul 2ul 4ul 8ul
Reaction program:
95c 10min -> 63C 14h -> add 1ul SLN mix(2U/ul AmpliTaq Stoffel fragment; 0.5U/ul AmpLigase; 0.8nM dNTP) -> 60C 4h ->
12 cycels (95C 1min -> 0.1C/sec to 60C, 60C for 4h 55C 1h) -> 95C 1min -> 37C 1min ->
add 2ul Exonuclease I/III mix -> 37C 2h -> 94C 5min -> 4C hold.
PCR[edit]
x7 Template 10ul 2X iProof Mastermix 50ul 350ul Amp_F2_sol(10uM)(with tail) 4ul 28ul Amp_R_sol (10uM)(with tail) 4ul 28ul 50X SYBG I 0.8ul 5.6ul H2O 31.2ul 218.4ul
98C 30S -> (98C 10S -> 58C 20S -> 72C 20S) x 8 -> (98C 10S -> 72C 20S) x 8 -> 72C 5 min -> 15C hold.
File:20080924 Yuan Gao sample SNP84k capture.jpg20080924_Yuan Gao sample SNP84k capture
Qiaquick column purification. Yields:
G2: 26.1ng/ul (132nM) x 30ul 2B: 26.8ng/ul (140nM) x 30ul 3B: 40.7ng/ul (213nM) x 30ul H1: 33.4ng/ul (175nM) x 30ul H2: 43.3ng/ul (227nM) x 30ul
Sent 15ul products to Yuan Gao on 09/24/08.