IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/08/24

iGEM Project name 1

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QS Track Like the dspB track, a bacterial mat grew when transformants were spread on chloramphenicol plates. A bacterial mat also grew when transformants were spread on LB + 20 μL chloramphenicol (1000X). This suggests a possible problem with the chloramphenicol aliquots (1000X) used. Marianne remade the chloramphenicol aliquots using 3 different sources of chloramphenicol. Phillip made chloramphenicol plates (20) using Marianne's C3 aliquots (chlor source: last year's parafilm sealed bottle) From yesterday's leftover ligation mixtures, 5 μL was transformed again by Vicki (following UBC iGEM procedures). Also started an overnight culture of pcN33. Host is Staph aureus RN4220. Used LB broth (no erm in lab). Made an antibiotic test of possible problematic chloramphenicol aliquots mentioned earlier. 10 μL of chloramphenicol aliquot was added to 10 mL LB. A stab of stock DH5(alpha)was inoculated in this medium. Expect no growth. dspB Track Gel verification on RD of chlor & amp backbones Protocol: SOP Changes: 0.08% agar Machine conditions: 0.5x TBE buffer, 100V, 60 min Gel orientation: Gel orientation 1 kb ladder Chlor E Chlor X Chlor P 100 bp ladder Amp E Amp X Amp P 1 kb ladder Results: Transformation on ligation mixes from Aug 13 2010 Ligation mixes: 1,4,5,8,11,14,15,AA,TB,his, no-his Protocol: Common protocol - SOP Changes: add 2uL of ligation mixes instead of 1uL Transformants put in 37C incubator @ 1630 O/N Culture for making competent cells Put in at 1630 Biofilm Track Eric F. and Melody Biofilm Growth Protocol Day 2 (100823E + M) O/N cultures removed from incubator @ 1030 - 20 hours in incubator Control showed no growth Plate Layout =100823E + M Row 3 4 5 6 7 8 9 10 C 8 C R C 8 C R C D 8 C R C 8 C R C R 8 C R C 8 C R C Note: R = RN4220 + 1% Glucose TSB 8 = 8325 - 4 + 1% Glucose TSB C = control + 1% Glucose TSB Any contamination that occurs in C8 (100823E) is isolated 100823E into incubator @ 1056 100823M into incubator @ 1305 Biofilm Growth Protocol Day 1 (100824E + M) Followed protocol RN4220 + 8325-4 + Control 5mL TSB each Into incubator @ 1500