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From 2012.igem.org

August 2

Mutation of FT

Inverse PCR

10xBufer	2mM dNTPs	primer fwd	primer rev	template	polymerase	MilliQ	Total
5	5	1.5	1.5	0.5	1	35.5	50

94°C

Dpn1 Digestion

PCR product	Dpn1
50	2

37℃,1h incubate

Self-ligation

product	MilliQ	Ligase	T4 Kinase	Total
2	7	5	1	15

August 10

Golden Gate Assembly method This method helps us to constract some genes quickly and we can design the order of constractions. We are trying to construct a gene cycle composed of 6 genes( pSB1A3, lacI, GFP, RFP, GFP, DT). After this experiment is confirmed to work, we will so some experiment to check how the numbers of promorters influences the strength of transcription.

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