Team:Paris Saclay/Notebook/August/9

From 2013.igem.org

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Objective : Obtaining FNR and BphR2 proteins (Gibson assembly)
Objective : Obtaining FNR and BphR2 proteins (Gibson assembly)
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===='''1 - Gibson assembly.'''====
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(((((((((((((((===='''1 - Gibson assembly.'''====
August 1st PCR purification to be sure about the experiment.  
August 1st PCR purification to be sure about the experiment.  
BphR2 part1, BphR2 part2 and RBS_BphR2_part1, FNR part1, FNR part2 and RBS_FNR_part1.
BphR2 part1, BphR2 part2 and RBS_BphR2_part1, FNR part1, FNR part2 and RBS_FNR_part1.
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we have no fragment so we must do again these PCRs  
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we have no fragment so we must do again these PCRs )))))))))))))))))
{{Team:Paris_Saclay/incl_fin}}
{{Team:Paris_Saclay/incl_fin}}

Revision as of 18:11, 16 September 2013

Navigation Home Team Project Labwork Notebook Human practices

Contents

Notebook : August 9

Lab work

A - Aerobic/Anaerobic regulation system

((((((((((((((((((((====1 - Obtaining Δ fnr E. coli strain by transduction to test our biobricks====

Abdou, Anais, Damir, Nadia, XiaoJing

We do the exprience again because the lysis made on wensday didn't happen. It's probably because the phage strain was too old ( 2001)

Protocol : transduction

Ps908transduction.jpg

Picture: lysed cells comparison.

  • 0µl phage(control): the petri dish is cloudy, bacteria are not lysed.
  • 50µl phage: the petri dish is clear, bacteria are lysed by phages.
  • We used a wild type strain to keep a stock and a Δ fnr E. coli strain to obtain phages that might have encapsidated a Δfnr::Km fragment.

10µl,50µl and 100µl petri dishes are clear so phages are multiplied.

We let the antibiotic over night to select the right strain. ))))))))))))))))))))))


Objective : obtaining Bba_K1155007

1 - Extraction of Bba_K115007 from DH5α

Abdou

Protocol : Hight copy plamid extraction

We used colonies number 10, 14 and 15.

Nanodrop

  • Bba_K1155007 in clone 10 : 38ng/µl
  • Bba_K1155007 in clone 14: 48.5ng/µl
  • Bba_K1155007 in clone 15: 52 ng/µl

CONCLUSION !!!!!!!! We will sequence our plasmid.


A - Aerobic/Anaerobic regulation system / B - PCB sensing system

Objective : Obtaining FNR and BphR2 proteins (Gibson assembly)

(((((((((((((((====1 - Gibson assembly.==== August 1st PCR purification to be sure about the experiment. BphR2 part1, BphR2 part2 and RBS_BphR2_part1, FNR part1, FNR part2 and RBS_FNR_part1.

Protocol : PCR_clean_up

Results:

IMAGE
  • Well 1 : 6µL DNA Ladder
  • Well 2 : 5µL of BphR2 part1+1µl of 6X loading dy
  • Well 3 : 5µL of BphR2 part2+1µl of 6X loading dy
  • Well 4 : 5µL of RBS_BphR2 part1+1µl of 6X loading dy
  • Well 5 : 5µL of FNR part1+1µl of 6X loading dy
  • Well 6 : 5µL of FRN part2+1µl of 6X loading dy
  • Well 7: 5µL of RBS_FNR part1+1µl of 6X loading dy
  • Gel : 0.8%

we have no fragment so we must do again these PCRs )))))))))))))))))

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