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Team:Hawaii/Biobricks extraction & transformation competency test
From 2008.igem.org
Contents |
Question
- Is the small amount of transformants we got on our "Initial BioBrick Plasmid Extraction From Filter Paper" experiment due to the low concentration of DNA from the filter paper or the low competency of our cells?
- How competent are our DH5α cells compared to commercial quality competent cells?
Protocol
The same protocol as "Initial BioBrick Plasmid Extraction From Filter Paper" was used.
- Filter paper was smashed up really well in 7 μl TE.
- Competency of our DH5α cells was compared to StrataCloneTM competent cells.
- Concentration of DNA on filter paper was estimated by transforming with pUC18 (.1 ng/μg) and comparing colony counts
- Cells were shaken at 200rpm.
- 1/1 and 1/10 dilutions were plated.
Results
Row1: Super Competent Cells
Row2: DH5α
L->R pUC18 1:1,pUC18 1:10, BBa_B0034 1:1, BBa_B0034 1:10
| Type of Cell | Plasmid | Dilution | Colonies |
| Strataclone Compentent Cells | pUC18 | 1:1 | Too many to Count |
| 1:10 | 232 x 4 | ||
| BBa_B0034 | 1:1 | 20 | |
| 1:10 | 1 | ||
| DH5α | pUC18 | 1:1 | Too many to Count |
| 1:10 | 43 | ||
| BBa_B0034 | 1:1 | 1 | |
| 1:10 | 1 |
Discussion
The StrataClone cells were much, much more competent than our cells. We should make more competent, competent cells if we have any chance of getting good transformants for our Biobricks parts and isolated genes.
