Team:Stockholm/5 October 2010

From 2010.igem.org

(Difference between revisions)
(→Andreas)
(→Andreas)
Line 29: Line 29:
#D
#D
#B & C
#B & C
 +
 +
===Transfer of nCPP⋅SOD⋅His.RBS.yCCS operon to pEX===
 +
====Plasmid preps====
 +
''From 4/10 ON cultures''
 +
 +
As I'm still waiting for sequencing results of the operons in pSB1K3, I spun down cells (4,400 x ''g'', 10 min), and saved pellets in -20 °C ON.
 +
 +
* pEX.nTAT⋅SOD⋅His.RBS.yCCS 2: A
 +
* pEX.nTAT⋅SOD⋅His.RBS.yCCS 3: B
 +
* pEX.nTra10⋅SOD⋅His.RBS.yCCS 1: B
 +
* pEX.nTra10⋅SOD⋅His.RBS.yCCS 2: B
 +
* pEX.nLMWP⋅SOD⋅His.RBS.yCCS 2: B
 +
* pEX.nLMWP⋅SOD⋅His.RBS.yCCS 3: B

Revision as of 14:31, 5 October 2010

Home Project Idea Lab Work Results Modelling Team       Follow us on and        2010.igem.org

↳   Notebook Safety Protocols


Contents

Andreas

Transfer of pEX.nCPP⋅SOD⋅His to BL21

Glycerol stocks

  • BL21 pEX.nLMWP⋅SOD⋅His
  • BL21 pEX.nTra10⋅SOD⋅His

Assembly of His⋅SOD⋅cCPP constructs

Colony PCR

  1. pSB1C3.His⋅SOD⋅cTra10 A-D
  2. pSB1C3.His⋅SOD⋅cTAT A-D
  3. pSB1C3.His⋅SOD⋅cLMWP A-D

Standard colony PCR settings

  • Elongation time: 1:30

Gel verification

Colony PCR gel verification of His⋅SOD⋅cCPP constructs in pSB1C3.
4 μl λ; 5 μl sample.
λ = O'GeneRuler 1 kb DNA ladder.

1 % agarose, 120 V

Expected bands

  1. 884 bp
  2. 854 bp
  3. 863 bp

Results
Varying band sized, with the following clones seeming correct:

  1. A
  2. D
  3. B & C

Transfer of nCPP⋅SOD⋅His.RBS.yCCS operon to pEX

Plasmid preps

From 4/10 ON cultures

As I'm still waiting for sequencing results of the operons in pSB1K3, I spun down cells (4,400 x g, 10 min), and saved pellets in -20 °C ON.

  • pEX.nTAT⋅SOD⋅His.RBS.yCCS 2: A
  • pEX.nTAT⋅SOD⋅His.RBS.yCCS 3: B
  • pEX.nTra10⋅SOD⋅His.RBS.yCCS 1: B
  • pEX.nTra10⋅SOD⋅His.RBS.yCCS 2: B
  • pEX.nLMWP⋅SOD⋅His.RBS.yCCS 2: B
  • pEX.nLMWP⋅SOD⋅His.RBS.yCCS 3: B
Retrieved from "http://2010.igem.org/Team:Stockholm/5_October_2010"