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Revision as of 13:42, 26 July 2010 (view source) Deenatsu (Talk | contribs) (→Conditions for ThermoCycler:) ← Older edit		Revision as of 13:43, 26 July 2010 (view source) Deenatsu (Talk | contribs) Newer edit →
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	# 2.5µl backward primer		# 2.5µl backward primer
	# 1µl template DNA		# 1µl template DNA
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	====Conditions for ThermoCycler:====		====Conditions for ThermoCycler:====

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Revision as of 13:43, 26 July 2010

Colony PCR

Materials to add accordingly:

1. 37.5µl of distilled H2O
2. 10µl of 5x GoTaq Buffer
3. Nucleotide DNTP
4. 2.5µl forward primer
5. 2.5µl backward primer
6. 1µl template DNA

Conditions for ThermoCycler:

1. Initialise - 95°C for 2 minutes.
2. Denature - 95°C for 30 seconds.
3. Anneal - 52°C for 30 seconds (Melting temperature, Tm of template).
4. Extension - 75°C for 30 seconds
5. Extension finish - 75°C for 5 minutes
6. Hold - 4°C

Steps 2 to 4 are repeated for 30 cycles before continuing to step 5.