IGEM:Peking University/2008/Experiment/PKU 0620 G2 1

Primer design

Primer 1

• Primer name: F-HindIII-Apal-Gal4
• Sequence(5'to3'):CCCAAGCTTGGGCCCAAGATGAAGCTACTGTCTTCTATCGAACAA
• Tm:64.9°C

Primer 2

• Primer name: R-EcoRI-Spel-Gal4
• Sequence(5'to3'):GGGGAATTCACTAGTATTTTACTCTTTTTTTGGGTTTGGTGGGGT
• Tm:61.3°C

PCR

TITLE: PCR-Budding Yeast Genome-Gal4-phusion polymerase 50*6ul

	Volume (μL)
Template （pPT1）	0.5
dNTP	1
Enzyme	0.5
Buffer	10
F-HindIII-Apal-Gal4	0.5
R-EcoRI-Spel-Gal4	0.5
ddH2O	37

conditions

Temperature	Time Length(0:00:00)
98°C	00:30
98°C	00:10	30 cycles
57.1,60.4,62.5,64.7,66.9,68.9°C	00:30	30 cycles
72°C	01:30	30 cycles
72°C	07:00
4°C	Hold

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Enzyme cutting-pPT1-XbaI-20 μL

	Volumn(μL)
plasmid	5
XbaI	0.2
BSA	2
M Buffer	2
ddH2O	10.8

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