User:Mbennie/Notebook/Lab Notebook/Notebook/2007/08/11

Cellular Adhesion

• Glycerol
  • Took 1.6ml of liquid culture and created 10% glycerol stocks
    • IgAbc #8

• Miniprep
  • Extracted DNA from remaining liquid culture (~6ml)
  • Eluted in 50ul water

• Liquid culture
  • Grew up cultures in 8ml Amp/Kan LB (didn't have mixed, so used 4ml of LB KAN and 4ml of LB Amp) inoculated from plates
    • IgAss + Fos #1
    • IgAss + Fos #2
    • IgAss + Fos #3
    • IgAss + JunB #1
    • IgAss + JunB #2
    • IgAss + JunB #3
  • Grew up cultures in 8ml Amp/Cl LB inoculated from plates
    • GCN4 #4
    • GCN4 #5
    • GCN4 #6

• PCR
  • Template: 40ul PCR Supermix, .4ul N. gonorrhea DNA, and .4ul of each primer
    • B: Part 1 of IgA beta-core IgAb-F and Mut_Pst1a-R
    • C: Part 2 of IgA beta-core Mut_Pst1a-F and Mut_Pst1b-R
    • D: Part 3 of IgA beta-core Mut_Pst1b-F and IgAb-R
    • E: Full IgA beta-core IgAb-F and IgAb-R
  • Same thermocycler protocol as 8.1.2007

• PCR
  • Template: 40ul PCR Supermix, .4ul VF2 and VR, 1ul IgAbc #8 miniprep
  • Same thermocycler protocol as 8.1.2007

Full IgA beta-core without muts, IgAbc #8, log-2 ladder

• Gel
  • IgAbc #8 looks correct
  • Try sequencing it again