Wisconsin-Madison/28 August 2009

August 28 2009
cloning with two single digests:

ProU PCR product with RBS/RFP

ProU PCR product with PSB1A2

ProU PCR product with GsSDMT

NudF with PSB1A2

Sequential Digestion

First Digestion

50uL / DNA / Buffer / H2O / Enzyme

ProU / 40 / 5 / 4 / EcoR I 1

nudF / 40 / 5 / 4 / XbaI 1.5

metK / 40 / 5 / 4 / XbaI 1.5

Hawaii / 40 / 5 / 4 / XbaI 1.5

RFP / 40 / 5 / 4 / EcoR I 1

Gssdmt / 40 / 5 / 4 / EcoR I 1

pSB1A2 / 40 / 5 / 4 / XbaI 1.5

PCR clean-up

Second Digestion

50uL / DNA / Buffer / H2O / Enzyme

ProU / 40 / 5 / 4 / Spe I 1

nudF / 40 / 5 / 4 / Pst 1.5

metK / 40 / 5 / 4 / Pst 1.5

Hawaii / 40 / 5 / 4 / Pst 1.5

RFP / 40 / 5 / 4 / XabI 1

Gssdmt / 40 / 5 / 4 / XbaI 1

pSB1A2 / 40 / 5 / 4 / PstI 1.5

Ran gel

Ligation