Team:Paris Saclay/Notebook/August/28

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Notebook : August 27

summary

  • We got clonies on ligation promoter fnr(activator)nirB plus RBS_LacZ+Term_PSB1C3

promoter fnr(activator)nirB plus RBS_AmilCP+Term_PSB1C3 promoter fnr(repressor) plus RBS_LacZ+Term_PSB1C3 promoter fnr(repressor) plus RBS_AmilCP+Term_PSB1C3t so we do Colony PCR for it

  • We didn't get clonies on Gibson so we do a gel electrophoresis of parts of Gibson assembly to verify the size
  • Digestion Dnp1 purification of the the PSB1C3 clean for Gibson and electrophoresis of it .


lab work


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

    • 1 - Gel extraction of the PSB1C3 Clean for Gibson.

Protocol : Gel extraction

    • 2- Migration of 3µl PSB1C3 Clean for Gibson (product gel extraction) '
[[]]
  • Well 1 : 6µL DNA Ladder
  • Well 2 : 3µL of PSB1C3 Clean dig Dnp1 gel extraction


  • Gel : 1.0%
    • 3- product quantification.
We used the nano drop to measure the DNA in 260nm and we found its concentration
name PSB1C3 Clean dig Dnp1 gel extraction
conc. 37.2ng/µl



    • 4 - Gibson assembly.


  • RBS_BphR2_part1, BphR2_part2 and plasmid PSB1C3
  • FNR_part1, FNR part1 and plasmid PSB1C3
  • RBS_FNR part1, FNR_part2 and plasmid PSB1C3


Sample Volume:

  • Tube 1 : RBS_BphR2 part1(1ul), BphR2 part2(1ul) , plasmid PSB1C3(3ul),MIX Gibson (15 ul)
  • Tube 2 : FNR part1(1ul), FNR part2(1ul) , plasmid PSB1C3(3ul),MIX Gibson (15 ul)
  • Tube 3 : RBS_FNR part1(1ul), FNR part2(1ul) , plasmid PSB1C3(3ul),MIX Gibson (15 ul)

These 3 mixture is incubated at 50°C for up to one hour. Transformation and spread in 6 LB plates with Chlorenphenicol . Incubate at 37°C over night.

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