Notebook : July 3

Lab work

A - Aerobic/Anaerobic regulation system

Objective : obtaining BBa_K1155000

1 - Colony PCR of BBa_K1155000 to check good insertion of Pndh* in pSB1C3

Abdou, Sheng, Zhou

Colonies count for BBa_K1155000 :

• Standard concentration :
  • BBa_K1155000 : 0

• High concentration :
  • BBa_K1155000 : 2

Primers and PCR :

VF2, VR, Pfnr_up, Pfnr_down are four oligos that we used for plasmid amplification. We used tree combinaisons VF/VR, VF/Pfnr_Down, Pfnr_Up/VR. If the promoter Pfnr insert PSB1C3 plasmid successfully, tree fragments with specific size will be amplified.

We mix each colony with 20µL of H2O.

Used quantities :

• DNA : 2µL
• Mix : (it was divided in tubes for 4 different colonies for each oligo combinaison with 23µL of mix in each tube)
  • VF or Pfnr_Up : 6µL
  • VR or Pfnr_Down or VR : 6µL
  • dNTP : 6µL
  • Buffer Dream Taq : 30µL
  • Dream Taq : 6µL
  • H2O : 246µL

2 - Culture of BBa_K1155000

Sheng

We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_K1155000. We also did liquid culture of each one. We incubate culture at 37°C.

Objective : obtaining BBa_K1155003, BBa_K1155007

1 - Culture of BBa_I732017, BBa_K592009

Sheng

We made new cultures by streaking 2 colonies of 07/02/13 culture of BBa_I732017, BBa_K592009. We also did liquid culture of each one. We incubate culture at 37°C.