User:Sean P Corum/Notebook/PHIX174 Cell Free/2012/09/14: Difference between revisions
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** 6.25 μL 2mM dNTPs (each) (0.25mM each final) | ** 6.25 μL 2mM dNTPs (each) (0.25mM each final) | ||
** 5 μL 10X reaction buffer | ** 5 μL 10X reaction buffer | ||
** 1.56 μL 3.2 nM ΦX174 template (0.1 nM final) | ** 1.56 μL 3.2 nM ΦX174 template (0.1 nM final) | ||
** 1 μL PfuUltra II fusion HS DNA polymerase | ** 1 μL PfuUltra II fusion HS DNA polymerase | ||
* Aliquot 2 × 19.6 μL, and add: | * Aliquot 2 × 19.6 μL, and add: | ||
** 0.4 μL water (to be 1-2) | ** 0.4 μL water (to be 1-2) | ||
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* Next on the list: | * Next on the list: | ||
*# DpnI digestion (w/ and w/o PCR purification) | *# DpnI digestion (w/ and w/o PCR purification) | ||
*# Topisomerase IV / Gyrase (preceded by PCR purification) - how to assay linking number? | *# Topisomerase IV / Gyrase (preceded by PCR purification) - how to assay linking number? |
Revision as of 16:03, 14 September 2012
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Hypothesis 2: Gene L is necessary for phage propagation.
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