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Team:University of Ottawa/17 July 2008
From 2008.igem.org
Contents |
Today in the Lab
Chris
- Purification of AtCRE
- used PCR clean up kit to purify AtCRE sample
- Tranformation of Competent Cells
- followed transformation of competent cells protocol to insert AtCRE plasmid into preprepared competent E. coli cells.
- allowed cells to incubate overnight at 37 C
Matt
- Ligation
- As suggested by Dan the ligation was spiked with 1 ul ATP.
- I then performed a PCR cleanup of the Ligation product in order to purify it for transformation into competent cells.
- Transformation
- A transformation was performed on PTP2 ligation product - I then left the streaked plates to incubate for tomorrow.
- A glycerol stock was finally made of the BY4642 yeast strain int. DQ232597.
Dan
- Gel of construct 1 after ligation
- Gel was unsucessful, the wrong primers were used for PCR amplification.
- Ligation seemed to work very efficiently, digesting in 50 uL with the new ClaI enzyme is working very well.
- Digestions
- Three digestions were performed 1+T, 1+T (higher concentration of vector and insert), and just the vector
- Ligations
- Three above plasmids were ligated overnight
Tammy
- Plasmid DNA Isolation
- XL10 Competent E.Coli cells transformed with pDR197::AtCKX2 were lysed and plasmid DNA was isolated using the Sigma-Aldrich kit.
- Average DNA concentration approximately 80 ng/μL
- Digestion of isolated pDR197::AtCKX2
| Reaction Components | 1X V (μL) |
| H2O | 6 |
| Buffer 3 | 1 |
| BSA | 1 |
| BamHI | 0.05 |
| PstI | 0.05 |
| DNA | 2 |
| Total | 10 |
- Control - No DNA (H2O)
- PURE I
- 1:10 II
- 1:100 I
- 1:100 II
