Paris/July 31
From 2007.igem.org
< Paris
pTOPO cloning
Cloning of PCR products: P5, P6, P7, P8, P9, P10 (see Paris/Freezer)
Transformation
Ligations from yesterday
PCR mutagenesis DGAT
DGAT has Pst1 restriction site. We want to mutagenize it to do a biobrick.
| PCR : DGAT1 | ||||||
|---|---|---|---|---|---|---|
| PCR Settings | Buffer (5x) | 5x 10µL | Expected size | |||
| Annealing (°C) | MgCl2 10µM | 10µM 0µL | 1087 | |||
| 50 | dNTP 10µM | 10µM 1µL | Success | |||
| Time Elongation | Oligo F 10µM | 27 ForDGAT1 | 2.5µL | YES | ||
| 2m00' | Oligo R 10µM | 13 DPst1-DGAT-R | 2.5µL | Image (click to enlarge) | ||
| Number cycles | Water | 33µl | 
| |||
| 35 | Polymerase | Phusion 0.5µL | Band (0=ladder) | |||
| DNA | Miniprep pKS::DGAT (1µL = 138ng) | 3, 4, 5, 6 (11 = control without DNA matrix) | ||||
| PCR : DGAT2 | ||||||
|---|---|---|---|---|---|---|
| PCR Settings | Buffer (5x) | 5x 10µL | Expected size | |||
| Annealing (°C) | MgCl2 10µM | 10µM 0µL | 396 | |||
| 50 | dNTP 10µM | 10µM 1µL | Success | |||
| Time Elongation | Oligo F 10µM | 12 DPst1-DGAT | 2.5µL | YES | ||
| 2m00' | Oligo R 10µM | 27 Rev-DGAT1 | 2.5µL | Image (click to enlarge) | ||
| Number cycles | Water | 33µl |
| |||
| 35 | Polymerase | Phusion 0.5µL | Band (0=ladder) | |||
| DNA | Miniprep pKS::DGAT (1µL = 138ng) | 7,8,9,10 (11 = control without DNA matrix) | ||||
