Team:Hawaii/Notebook/2008-10- 8

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Things we did today

Wetlab work

Triparental conjugation (cont.)

Grace
  • PCR'd BBpRL cultures to ensure that plasmid was retained
  • Plasmid was not retained (grown in SOB w/o selection = no reason to retain plasmid)
  • PCR'd BBpRL colonies #1 and 18 from plate to ensure plasmid present
  • Regrew RP1, BBpRL harboring cultures in 20ml SOB with selection (Kan50 and Sp100 respectively) and incubated at 37C overnight, with shaking (rpm = 250)


Verification of transformants

Grace
EtBr stained 25 agarose gel ran at 95V for 1 hour. Two microliters of PCR product were loaded into each well.
Construct Colonies
nir + rgt#1 1
plac + rgt#1 0
nr + sg 12
pr + sg 74
nr + pgt 5
pr + pgt 19
pSB1A3 (control) 1
no DNA 0
  • Colony PCR of transformants
  • Restreaked nrsg #6, prpgt #5, 7, 11

Construction of secretion device (cont.)

Grace
  • PCR:
  • J33207
  • nir+rbs
  • nir
  • rgt#1
  • Overnight RE digest:
  • HindIII/BamHI:
  • J33207
  • EcoRI/SpeI:
  • nir
  • nir+rbs
  • XbaI/PstI:
  • rgt#1

Drylab work

iGEM title/abstract

Grace & Margaret<strong>
  • Revised abstract
  • Sent in abstract, title, track selection to iGEM HQ

Discussion

Quote of the Day

History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson


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