Team:Paris/19 August 2009
From 2009.igem.org
NoteBook
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Lab work
Sequencing w/ Bobode
5 clones pLac/RBS-TetR have to be sequenced
Clone 1, 2, 6, 7, 8 used (because of their high DNA concentration)
Transformation w/ Bobode
pSB1A3-TolRII ligation was transformed into P10 chemical competent cells, using the "heat shock" protocol
Glycerol stock w/ Charlie
Glycerol stock of :
S39 - pINTE3 (Col E3, AmpR)
S40 - pINTg3p (g3p Term, AmpR)
S41 - pINK2(TolRII, AmpR)
S42 - pTA (pTet-Col A, AmpR)
A28 OmpA signal (ex A11)
A29 FecR mutant
PCR 2 steps : 72°C 30s elongation
Gel verification Ok (OmpAsignal: weak)
Purification Ok
OD : A28=0,82 A29=0,92
Miniprep send to sequencing.
Miniprep DO: ClyA colonie 3:1,40
ClyA colonie 7:1,60
ClyA colonie 8:0,8
ClyA colonie 10:1,65
The results were transformed into µg/mL
Mix for sequencing:
Cly A colonie 3 | Cly A colonie 7 | Cly A colonie 8 | Cly A colonie 10
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Miniprep | 4,29µL | 3,75µL | 7,50µL | 3,64µL
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Oligos | 2,45µL | 2,45µL | 2,45µL | 2,45µL |
H2O | 8,26µL | 8,80µL | 5,05µL | 8,91µL |