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From 2010.igem.org

Contents 1 Mimmi 1.1 MITF-m 1.1.1 Gel 1.1.2 Re-ligation and transformation

Mimmi

MITF-m

Gel

well	sample
1	ladder
2	pSB1C3.MITF-M 8
3	pSB1C3.MITF-M 9
4	pSB1C3.MITF-M 10
5	pSB1C3.MITF-M 11
6	pSB1C3.MITF-M 12
7	pSB1C3.MITF-M 13
8	pSB1C3.MITF-M 14
9	positive control
10	digested MITF (E+S)
11	digested pSB1C3 (E+S)

Re-ligation and transformation

• Depohosphorylate vector:

Mix			Conditions
cut pSB1C3	9		time	°C
A phosp	1		10m	37
tot	10µl		5m	75
			oo	10

• Ligate

Mix	(µl)		Conditions
cut pSB1C3	2		time	°C
cut MITF-M	13		10m	22
5x buffer	4
T4 ligase	1
sH2O	0
tot	20µl

• Follow the original transformation protocol