TAIL PCR反应体系和程序
TAIL-PCR 反应体系(20μl体系)
Templet 1μl
10x buffter 2.0μl
Mg 2+ 1.5μl
dNTP 0.2μl (10mmol)
P1 0.2μl (15μmol)
P2(AD) 1.3μl (30μmol)
Taq 0.2μl (5U/μl)
ddH 2 O 15.1μl
Tail-PCR 反应程序及条件
Program used for Tail-PCR amplification
|
第一次扩增 1st amplification |
第二次扩增 2ed amplification |
第三次扩增 3rd amplification |
|
92 ℃3min |
94 ℃30min ＊＊＊ |
94 ℃10s |
|
95 ℃1min |
58 ℃1min ＊＊＊ |
94 ℃30s ＊＊＊＊ |
|
94 ℃30 s ＊ |
72 ℃2min ＊＊＊ |
42 ℃30s ＊＊＊＊ |
|
62 ℃1min ＊ |
94 ℃30s ＊＊＊ |
72 ℃2min ＊＊＊＊ |
|
72 ℃2min ＊ |
58 ℃1min ＊＊＊ |
72 ℃5min |
|
94 ℃30s |
72 ℃2min ＊＊＊ |
|
|
25 ℃3min |
94 ℃30s ＊＊＊ |
|
|
Ramp to72℃over 3min |
44 ℃1min ＊＊＊ |
|
|
72 ℃2min |
72 ℃2min ＊＊＊ |
|
|
94 ℃30s ＊＊ |
72 ℃5min |
|
|
44 ℃1min ＊＊ |
||
|
72 ℃2min ＊＊ |
||
|
94 ℃30s ＊＊＊ |
||
|
62 ℃1min ＊＊＊ |
||
|
72 ℃2min ＊＊＊ |
||
|
94 ℃30s ＊＊＊ |
||
|
62 ℃2min ＊＊＊ |
||
|
72 ℃2min ＊＊＊ |
||
|
94 ℃30s ＊＊＊ |
||
|
44 ℃1min ＊＊＊ |
||
|
72 ℃2min ＊＊＊ 72 ℃5min |
||
注： ＊，＊＊，＊＊＊，＊＊＊＊分别代表x5 ，x10 ，x15 ，x30 循环
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