Team:Pasteur Paris/Test
Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
07.16.2018
We transformed pET 43.1a and pSB1C3 in DH5-α competent cells, in order to constitute a stock of empty vectors for our manipulation.
| Plasmid | C (ng/μl) | Volume (μl) | Competent cell | Medium |
| pET 43.1.a | 4.95 | 1 | DH5-α | LB/carbenicilline |
| pSB1C3 | 40 | 1 | DH5-α | LB/chloramphenicol |
See here the transformation of E. coli DH5-alpha protocol We let the transformed bacteria grow overnight (16 hours).
07.17.2018
Results: We went to see our bacterial culture:
- Bacteria transformed with pET 43.1 had not grown.
- Bacteria transformed with pSB1C3 had formed colonies.
Interpretations: Bacteria transformed with pET 43.1 did not grow. We found a non-commercial tube of pET 43.1 in the freezer from last year team, and we decided to try to amplify it because we did not have any commercial tubes of pET 43.1. Transformation did not work as expected, probably because:
- There was no DNA left in the tube.
- The concentration of DNA was too low .
We cultivated the transformed pSB1C3 bacteria in liquid medium 2 x 25ml + Chloramphenicol (25µg/ml) overnight at 37°C, 180 rpm.
Results: Bacteria successfully transformed with pSB1C3. (See here the liquid culture protocol)
07.18.2018
Extraction: We extracted the pSB1C3 plasmid from the bacterial culture. The protocol used was the Qiagen Plasmid Purification Kit (See protocol here ) Measure of the DNA concentration in each tube thanks to the NanoDrop (Blank used : TE.1) (See the NanoDrop protocol here)
Results: We used the Nanodrop to quantify the purified DNA.
| Sample | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 |
| Volume (µl) | 50 | 50 | 50 | 50 | 50 | 50 | 50 | 50 | 50 | 50 |
| C (ng/µl) | -7.8 | -8.5 | 54.8 | 5.6 | 237.4 | -7.9 | -7.2 | -7.5 | -6.9 | -6.5 |
| 260/280 | 1.43 | 1.42 | 1.97 | 3.04 | 1.85 | 1.32 | 1.47 | 1.56 | 1.35 | 1.44 |
Remarque: [NA] < 0 = no DNA [NA] > 200 good 260/280 [1;2]
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
Something was done one that day
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
Something was done one that day
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
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Bacteriology
Cell culture
Microfluidics/Membrane
Product Design
Something was done one that day
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