Team:Groningen/protocols/PCR

From 2013.igem.org

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Revision as of 10:52, 27 July 2013

PCR

Materials:

  • PCR tubes
  • F- and R-primer
  • DNTPs
  • Phusion buffer
  • MQ water
  • Phusion Polymerase
  • DNA template

Steps:

Component 50μl reaction Final concentration
MQ water up to 50μl
5x Phusion Buffer 10μl 1x
10mM dNTPs 1μl 200μM
F-primer 2.5μl 0.2μM
R-primer 2.5μl 0.2μM
Template DNA 1μl ~1ng
Phusion Pol. 2U/μl 0.3μl 0.02U/μl
DMSO 5% 1.5μl

Cycling instructions:

Temperature Time Number of cycles
98°C 30sec 1
98°C 10sec 30
primer annealing temperature 30sec
72°C 30sec/kb
72°C 10min 1
4°C ∞ 1

Reference: http://www.thermoscientificbio.com/uploadedFiles/Resources/f-530s-product-information.pdf