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Team:Paris Saclay/Notebook/July/18
From 2014.igem.org
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[https://2014.igem.org/Team:Paris_Saclay/Protocols/Extraction_of_the_Genomic_DNA_from_Bacteria_by_using_NucleoSpin®_Tissue Protocol] | [https://2014.igem.org/Team:Paris_Saclay/Protocols/Extraction_of_the_Genomic_DNA_from_Bacteria_by_using_NucleoSpin®_Tissue Protocol] | ||
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====Preparation of electrocompetent DY330 and transformation via pJBEI6409==== | ====Preparation of electrocompetent DY330 and transformation via pJBEI6409==== | ||
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# Spread on nine petri dishes (LB+Cm): 50 µL from each cuvette, 100 µL from each cuvette, remainder from each cuvette. Incubate overnight at 30°C. NB: before spreading the remainder, centrifuge it, then remove 750 µL of supernatant. | # Spread on nine petri dishes (LB+Cm): 50 µL from each cuvette, 100 µL from each cuvette, remainder from each cuvette. Incubate overnight at 30°C. NB: before spreading the remainder, centrifuge it, then remove 750 µL of supernatant. | ||
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==Photo of the Day== | ==Photo of the Day== | ||
[[File:Paris Saclay 18_july.jpg|600px|center]] | [[File:Paris Saclay 18_july.jpg|600px|center]] | ||
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| + | '''Members present''': | ||
| + | * Instructors and advisors: Solenne. | ||
| + | * Students: Arnaud, Fabio, Mathias, Romain and Sean. | ||
{{Team:Paris_Saclay/notebook_footer}} | {{Team:Paris_Saclay/notebook_footer}} | ||
Latest revision as of 15:40, 14 October 2014
Contents |
Friday 18th July
Lab work
The chassis coli Odor free
1 - Transformation of CaCl2 supercompetent cells
by Arnaud & Romain
Strains transformed: MG1655, MG1655Z1. from Bacterial Cultures made the 17th July
2 - Plasmid DNA Purification
by Fabio
- BBa_J23119 (x2)
from Bacterial Culture made the 17th July
Lemon scent
Preparation of electrocompetent DY330 and transformation via pJBEI6409
by Sean
A. Preparation of DY330 strain
Protocol
- Dilute 1 mL of DY330 in 100mL of LB at 30°C.
- When optical density is around .6, divide the 100mL into four 50mL centrifuge tubes. Centrifuge for four minutes at 4°C, 4000rpm. Discard supernatant.
- Repeat, but this time with 25mL of 10% glycerol solution in each tube.
- Repeat, but this time with 12.5mL.
- Remove supernatant and resuspend pellet with any remaining supernatant. Collect all of the strain from the four tubes then store in ice.
B. Transformation by electroporation
Protocol
- Fill three electroporation cuvettes as follows. a)50µL of DY330+2µL of pJBEI6409 plasmid. b)50µL of DY330+4µL of pJBEI6409. c)Control without DNA.
- Electroporation at 2500V, 132W, 40µF.
- Add 950 µL of cold LB in each cuvette.
- Incubate at 30°C and under agitation for two hours.
- Spread on nine petri dishes (LB+Cm): 50 µL from each cuvette, 100 µL from each cuvette, remainder from each cuvette. Incubate overnight at 30°C. NB: before spreading the remainder, centrifuge it, then remove 750 µL of supernatant.
Photo of the Day
Members present:
- Instructors and advisors: Solenne.
- Students: Arnaud, Fabio, Mathias, Romain and Sean.
