August/21 August 2009

From 2009.igem.org

transformation

to get new plasmid

⑤(RBS+rhlR+terminator+promoter(reg))

⑦（RBS＋cinR)

⑧（RBS＋luxI)

check the result of transformation

Checked the cell cultures transformed and plated out yesterday (8/10) for colony formation and made an approximate count of the number of colonies.

(plate number)-(location on plate) (no. of colonies)

            2-8G                         4
            2-21F                        0    
            2-6O                         2
            2-13N                       <15
             ⑤                          0

restriction and digestion

K204009

Vector		Insert
terminater(1-23L)	10	①-3(RBS+LasR)	12
EcoRI	1	EcoRI	1
XbaI	1	SpeI	1
No.2	2	No.2	2
dH₂O	4	dH₂O	4
total	20uL	total	20uL

K204010

Vector		Insert
RBS(1-2M)	10	EpsE+terminater(②-A)	12
SpeI	1	XbaI	1
PstI	1	PstI	1
No.2	2	No.2	2
dH₂O	6	dH₂O	4
total	20uL	total	20uL

K204011

Vector		Insert
p(tetR)(1-6I)	10	RBS+GYP+terminater(1-12M)	12
SpeI	1	XbaI	1
PstI	1	PstI	1
No.2	2	No.2	2
dH₂O	6	dH₂O	4
total	20uL	total	20uL

↓
37°C, 2hr

↓
gel electrophoresis
gel cut

'8/22
purification by [QIAquick Nucleotide Removal Kit]
↓
ligation

ligation
DNA        44
10* buffer 5
ligation   1
total      50uL
↓
16°C , 4hr

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