Haynes Lab:Notebook/Inteins/2013/03/01

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(Entry title)
Current revision (17:00, 5 March 2013) (view source)
(Entry title)
 
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*Made a brand new gel  
*Made a brand new gel  
1KBT  KAH9(186)  KAH9(160)  KAH10 (116)  KAH10 (106)   
1KBT  KAH9(186)  KAH9(160)  KAH10 (116)  KAH10 (106)   
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*Put 10 of ladder, 30 in other wells.  
+
*Put 10 of ladder, 30 in other wells.
 +
http://openwetware.org/wiki/Image:3-1-2013_gel.jpg
*Run dna recovery kit  
*Run dna recovery kit  
**Melted @ 55.6 degrees C for 6 min., vortex in between
**Melted @ 55.6 degrees C for 6 min., vortex in between
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*Left ligations for at least 30 mins.  
*Left ligations for at least 30 mins.  
*Add 50 uL of competent cells  
*Add 50 uL of competent cells  
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  • Completed a restriction/digest from really good mini prep!
  • Used brand new enzymes for the restriction/ digest
  • Made a brand new gel

1KBT KAH9(186) KAH9(160) KAH10 (116) KAH10 (106)

  • Put 10 of ladder, 30 in other wells.

http://openwetware.org/wiki/Image:3-1-2013_gel.jpg

  • Run dna recovery kit
    • Melted @ 55.6 degrees C for 6 min., vortex in between
    • 8uL + 7 water to elute dna in last step
  • Completed ligation/transformation
  • Used both 2x and 10x buffers
  • Get competent cells and keep on ice
  • Used both buffers to see if lab shared stock Roche ligation buffer was contaminated.
  • Used brand new ligase and ligation buffer
  • Left ligations for at least 30 mins.
  • Add 50 uL of competent cells



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