User:Stuart McKellar/Notebook/Bird Sex Testing/2012/12/02
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Current revision (23:00, 1 December 2012) (view source) (→Entry title) |
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| - | == | + | ==Trying all loris with 65°C PCR - p2/p8 == |
| - | * | + | Aim: To see if at 65°C i can differentiate males from females. |
| + | |||
| + | Method: | ||
| + | Master mix for all 8 samples plus -ve control(10 units of MM): | ||
| + | *25ul buffer (10x) | ||
| + | *20ul dNTPs | ||
| + | *15ul MgCl2 | ||
| + | *1ul Taq | ||
| + | *25ul P2 primer (F) | ||
| + | *25ul P8 primer (R) | ||
| + | *129ul H<sub>2</sub>OH20 | ||
| + | |||
| + | Tubes labelled as follows: | ||
| + | *A:1SWWBD | ||
| + | *B:2SWWBD | ||
| + | *C:R5#9 | ||
| + | *D:R5#51 | ||
| + | *E:R5#52 | ||
| + | *F:R5#100 | ||
| + | *G:Pink Gold | ||
| + | *H:banjo | ||
| + | *-ve: 1ul water | ||
| + | |||
| + | Add 1ul of relevant DNA to each tube and 24ul of MM. | ||
| + | |||
| + | PCR: 94°C for 2m30s, 35 cycles of: 95°C 30s, 65°C for 30s, 72°C for 30s; final elongation 72°C for 5 mins( actually went 10 mins by accident). | ||
| + | |||
| + | results to be determined via agar gel electrophoresis on 1% gel stained using gelgreen. | ||
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Trying all loris with 65°C PCR - p2/p8Aim: To see if at 65°C i can differentiate males from females. Method: Master mix for all 8 samples plus -ve control(10 units of MM):
Tubes labelled as follows:
Add 1ul of relevant DNA to each tube and 24ul of MM. PCR: 94°C for 2m30s, 35 cycles of: 95°C 30s, 65°C for 30s, 72°C for 30s; final elongation 72°C for 5 mins( actually went 10 mins by accident). results to be determined via agar gel electrophoresis on 1% gel stained using gelgreen.
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