IGEM:Harvard/2008/Lab Notebooks/DailyBook/Week9/Chemical and Light: Difference between revisions
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===Gel 2: 12-22=== | ===Gel 2: 12-22=== | ||
[[Image:8-18_colpcr_ta2.jpg]] | [[Image:8-18_colpcr_ta2.jpg]] | ||
* #12, 14 picked for 5mL culture | |||
Lane 1: 1 kb ladder | Lane 1: 1 kb ladder | ||
Revision as of 17:44, 18 August 2008
105, mtrB Ligations/transformations results 8/18
Parts to ligated came from 8/17
We used a 2:1 molar ratio of insert to vector for all of the ligations. The ligations were all transformed into TOP10 cells.
| Plasmid | Number of colonies |
| P105+P1'BB | 2 |
| P105+P3'BB | 102 |
| P101+P105 | 0 |
| P101+P108 | 0 |
| mtrB(BB)+P108 | 20 |
| mtrB(BB)+P116 | 4 |
| mtrB(BB)+P117 | 13 |
In retrospect, it appears that all the ligations with mtrB are wrong, since the digest was actually wrong.
105 Colony PCR 08/18
The annealing temperature was 55 °C and the elongation time was 2'45" (we used Platinum Taq supermix).
Gel 1: 1-11
- #2, 5 picked for 5mL culture
Lane 1: 1 kb ladder
Lanes 2-3: P105+P1'BB
Lanes 4-12: P105+P1'BB
Gel 2: 12-22
- #12, 14 picked for 5mL culture
Lane 1: 1 kb ladder
Lanes 2-12: P105+P1'BB
Gel 3
Lane 1: 1 kb ladder
Lanes 2-12: P105+P1'BB
RE digests 45, 63, 97 08/18
We used the Fermantas enzymes and digested for 25'.
Lane 1: 1 kb ladder
Lane 2: P45 cut XP (876; 2079)
Lane 3: P63 cut EX (3284)
Lane 4: P97 cut SP (2091)
