Difference between revisions of "PrbbBB:colony pcr v1"

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(New page: Back to all protocols ==Overview== ==Materials== *96-well PCR plate *AmpliTaq DNA Polymerase 5 U/µl (Roche) *AmpliTaq Buffer 10 x *dNTP mix 10mM each nucleotide ...)
 
(Materials)
Line 12: Line 12:
 
*ddH2O
 
*ddH2O
 
*primers:
 
*primers:
**BBa_G00100 (VF2) [http://partsregistry.org/wiki/index.php/Part:BBa_G00100 MIT Registry] [http://brickit.crg.es/registry/biobrick/5/ CRG-internal registry]
+
**[http://partsregistry.org/wiki/index.php/Part:BBa_G00100 BBa_G00100 (VF2)] [http://brickit.crg.es/registry/biobrick/5/ CRG-internal registry]
**BBa_G00101 (VR)  [http://partsregistry.org/wiki/index.php/Part:BBa_G00101 MIT Registry] [http://brickit.crg.es/registry/biobrick/6/ CRG-internal registry]
+
**[http://partsregistry.org/wiki/index.php/Part:BBa_G00101 BBa_G00101 (VR)] [http://brickit.crg.es/registry/biobrick/6/ CRG-internal registry]
  
 
==Procedure==
 
==Procedure==

Revision as of 04:50, 26 February 2009

Back to all protocols

Overview

Materials

Procedure

II PCR reaction

100µl single reaction 3.5xMaster 10xMaster
H2O 76µl 266 760
5x HF Buffer 20µl 70 200
10mM dNTP 2µl 7 20
FW primer 100 µM 0.5µl 1.75 5
RV primer 100 µM 0.5µl 1.75 5
Phusion 1µl 3.5 10
template DNA 0.1µl -- --
PCR Program
30"@98°C
5x (10"@98°C; 15"@Ta; text@72°C);
25x (10"@98°C; text@72°C);
10'@72°C
∞ 4°C
  • extension time text = (kb insert length) × 25"
  • annealing temperature Ta = (primer annealing) + 3°C

II Agarose Gel

Notes

Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!

raik: no comment


References

Contact

or instead, discuss this protocol.