"
Team:Newcastle/24 August 2010
From 2010.igem.org
| |||||||||||||
| |||||||||||||
Contents |
Miniprep for pGFPrrnB with yneA
Aims
The aim of this experiment is to produce more stocks of yneA in pGFPrrnB.
Materials and Protocol
Please refer to qiagen minipreps and nanodrop spectrophotometer protocols.
Results
The results from the nanodrop:
| DNA | Concentration (ng/µl) |
|---|---|
| 1 | 286.1 |
| 2 | 304.0 |
| 3 | 316.3 |
| 4 | 421.6 |
| 5 | 518.7 |
| 6 | 460.1 |
| 7 | 370.1 |
| 8 | 377.3 |
| 9 | 346.0 |
| 10 | 347.4 |
| 11 | 202.8 |
| 12 | 307.4 |
Discussion
The results from the nanodrop showed that we have produced high concentration of vector pGFPrrnB with insert yneA. We will then proceed to digestion.
Single and Double Digestion of pGFPrrnB with yneA
Aims
To check if the insert yneA has been inserted into vector pGFPrrnB.
Materials and Protocol
We are doing two digests for pGFPrrnB and yneA:
- Single digest with HinDIII;
- Double digest with EcoR1 and Nhe1.
Please refer to restriction digests and gel electrophoresis.
Results
Gel electrophoresis results for digestion:
Discussion and Conclusion
Single digestion of pSB1C3
Aims
To linearise the vector pSB1C3. The linear vector will be used as a PCR template for Gibson cloning of the Subtilin immunity and rocF BioBricks.
Materials and Protocol
We are doing a single digest with HinDIII.
Please refer to restriction digests and gel electrophoresis.
Results
Gel electrophoresis results for digestion:
Figure 1: Gel electrophoresis of the amplified PCR products and restriction digest
- Lane 1: 1 Kb ladder
- Lane 2: digested (linearised) pSB1C3
Discussion and Conclusion
The digestion was successful — the correct band was seen. We can proceed to gel extraction.
   
|
    
|
