Team:NYU-AD/Notebook/Week6

Week 6

08/09/15

  • Miniprepped the liquid cultures from lldd_ IDT, 4 tubes.
  • Resuspended RFP in PCSB1C3 backbone (shipping plasmid) from distribution kit in 10uL nuclease-free water ( Plate 4, well 4B)
  • Transformed 2 tubes with RFP backbones to amplify, 4uL plasmid per tube.
  • Plated on 2 plates, 100uL mixture per plate. The plates were plates in the incubator
  • Also performed the digestion and ligation : Promoter + LLD on AmpR.
  • 4 separate tubes of ligation product, labelled P+LLD ligate 1-4 is in the freezer box
  • tubes 1 and 2 were ligated to Amna’s AmpRbackbone
  • tubes 3 and 4 were ligated to Amna’s AmpRbackone 2

09/09/2015

  • Transformed Pro +lldd contruct ( AmpR). The 4 plates are in the incubator
  • Miniprepped the Pro+ TnaA contruct ( KanR). The 6 tubes are in the freezer box, the average concentration is 20ng/uL

Digestion And Ligation

  • We followed the standard digestion and ligation protocol
  • Join Pr +TnaA to TnaB + Ter and put on both pSBC13 and AmpR backbone

    • PHOTO

  • Used tubes 4,5,6, from Pro+TnaA miniprep from 9/09
  • We halved the volumes of digestion procedure to give total digest volume of 25uL.

10/09/2015

  • Picked the colonies of the tnaA and lldd and inoculated the final construct on AmpR backbone and shipping plasmid

11/09/2015

  • Miniprepped the final construct on AmpR backbone and shipping plasmid

12/09/2015

  • Switched the final construct of tnaA and lldd onto shipping plasmid

Week 7

13/09/2015

  • Transformation of shipping constructs lldd and tnaA onto shipping plasmid using previous standard protocol
  • Created 4 replicates for each

14/09/2015

  • picked the white colonies and inoculated the liquid cultures following standard protocol

15/09/2015

  • miniprepped the final construct for shipping