Team:Paris Saclay/Notebook/July/18

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{{Team:Paris_Saclay/notebook_header}}
=Friday 18th July=
=Friday 18th July=
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==Lab work==
==Lab work==
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===The chassis coli Odor free===
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===1 - Transformation of supercompetent cells with CaCl2===
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====1 - Transformation of CaCl<sub>2</sub> supercompetent cells====
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''by Arnaud & Romain''
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''by Romain & Arnaud''
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Strains transformed: MG1655, MG1655Z1.
Strains transformed: MG1655, MG1655Z1.
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[https://2014.igem.org/Team:Paris_Saclay/Protocols/Transformation_of_supercompetent_Ecoli_cells_with_CaCl2 Protocol]
[https://2014.igem.org/Team:Paris_Saclay/Protocols/Transformation_of_supercompetent_Ecoli_cells_with_CaCl2 Protocol]
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===2 - Plasmid DNA Purification===
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====2 - Plasmid DNA Purification====
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''by Fabio''
''by Fabio''
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*J23119 (x2)
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*BBa_J23119 (x2)
from Bacterial Culture made the [https://2014.igem.org/Team:Paris_Saclay/Notebook/July/17 17th July]
from Bacterial Culture made the [https://2014.igem.org/Team:Paris_Saclay/Notebook/July/17 17th July]
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[https://2014.igem.org/Team:Paris_Saclay/Protocols/Extraction_of_the_Genomic_DNA_from_Bacteria_by_using_NucleoSpin®_Tissue Protocol]
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People there:
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===Lemon scent===
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====Preparation of electrocompetent DY330 and transformation via pJBEI6409====
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''by Sean''
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'''''A. Preparation of DY330 strain'''''
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Protocol
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# Dilute 1 mL of DY330 in 100mL of LB at 30°C.
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# When optical density is around .6, divide the 100mL into four 50mL centrifuge tubes. Centrifuge for four minutes at 4°C, 4000rpm. Discard supernatant.
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# Repeat, but this time with 25mL of 10% glycerol solution in each tube.
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# Repeat, but this time with 12.5mL.
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# Remove supernatant and resuspend pellet with any remaining supernatant. Collect all of the strain from the four tubes then store in ice.
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'''''B. Transformation by electroporation'''''
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Protocol
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# 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.
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# Electroporation at 2500V, 132W, 40µF.
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# Add 950 µL of cold LB in each cuvette.
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# Incubate at 30°C and under agitation for two hours.
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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.
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==Photo of the Day==
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[[File:Paris Saclay 18_july.jpg|600px|center]]
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'''Members present''':
* Instructors and advisors: Solenne.
* Instructors and advisors: Solenne.
* Students: Arnaud, Fabio, Mathias, Romain and Sean.
* Students: Arnaud, Fabio, Mathias, Romain and Sean.
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{{Team:Paris_Saclay/notebook_footer}}
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[https://2014.igem.org/Team:Paris_Saclay/Notebook Back to the calendar]
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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

Protocol

2 - Plasmid DNA Purification

by Fabio

  • BBa_J23119 (x2)

from Bacterial Culture made the 17th July

Protocol

Lemon scent

Preparation of electrocompetent DY330 and transformation via pJBEI6409

by Sean

A. Preparation of DY330 strain

Protocol

  1. Dilute 1 mL of DY330 in 100mL of LB at 30°C.
  2. When optical density is around .6, divide the 100mL into four 50mL centrifuge tubes. Centrifuge for four minutes at 4°C, 4000rpm. Discard supernatant.
  3. Repeat, but this time with 25mL of 10% glycerol solution in each tube.
  4. Repeat, but this time with 12.5mL.
  5. 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

  1. 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.
  2. Electroporation at 2500V, 132W, 40µF.
  3. Add 950 µL of cold LB in each cuvette.
  4. Incubate at 30°C and under agitation for two hours.
  5. 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

Paris Saclay 18 july.jpg

Members present:

  • Instructors and advisors: Solenne.
  • Students: Arnaud, Fabio, Mathias, Romain and Sean.

Back to the calendar

Retrieved from "http://2014.igem.org/Team:Paris_Saclay/Notebook/July/18"