Haynes Lab:Notebook/SynBERC Scholars 2013/2014/03/17: Difference between revisions

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|-
|-
| REAGENT || 1 || 2 || 3 || 4 || 5 || 6  
| REAGENT || 1 || 2 || 3 || 4 || 5 || 6  
| rowspan="6" | [[Image:PrimerCheck_031814.JPG|250px|Primer check gel]] <br>2% agarose gel
| rowspan="6" | Inconclusive: Gel showed no DNA smear
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| Genomic DNA (~500 ng) || 2.0 μL || 2.0 || 2.0 || 1.0 || 1.0 || 1.0
| Genomic DNA (~500 ng) || 2.0 μL || 2.0 || 2.0 || 1.0 || 1.0 || 1.0
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* Repeat PCR from [http://openwetware.org/wiki/Haynes_Lab:Notebook/SynBERC_Scholars_2013/2014/03/06 3/06/14]
* Repeat PCR from [http://openwetware.org/wiki/Haynes_Lab:Notebook/SynBERC_Scholars_2013/2014/03/06 3/06/14]
#




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| Reagent || Kozak || Luc14
| Reagent || Kozak || Luc14
| rowspan="6" | [[Image:PrimerCheck_031814.JPG|250px|Primer check gel]] <br>2% agarose gel
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| DNA || 0.5 μL || 0.5  
| DNA || 0.5 μL || 0.5  

Revision as of 15:44, 21 March 2014

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03/17/14

  • Luc 14 genomic DNA test - restriction digest and gel
  • Luc 14 genomic DNA clean-up & PCR


Luc 14 genomic DNA test - restriction digest and gel

  • Check genomic DNA on a gel, with or without restriction digestion
  1. Luc 14 HEK - No enzyme
  2. Luc 14 HEK - EcoRI (fast digest)
  3. Luc 14 HEK - PstI (fast digest)
  4. U2OS plain - No enzyme
  5. U2OS plain - EcoRI (fast digest)
  6. U2OS plain - PstI (fast digest)
REAGENT 1 2 3 4 5 6 Inconclusive: Gel showed no DNA smear
Genomic DNA (~500 ng) 2.0 μL 2.0 2.0 1.0 1.0 1.0
Rest. enzyme 0 2.0 2.0 0 2.0 2.0
10x buffer 2.0 2.0 2.0 2.0 2.0 2.0
dH2O 16.0 14.0 14.0 17.0 15.0 15.0
  20.0 20.0 20.0 20.0 20.0 20.0


Luc 14 genomic DNA clean-up & PCR

  • Clean up 1/2 of the Luc genomic DNA (50 μL) on a Zymo clean and concentrator column
    • 50 μL ~ 4340 ng
    • Eluted with 20 μL dH2O ~ 217 ng/μL


Reagent Kozak Luc14 Primer check gel
2% agarose gel
DNA 0.5 μL 0.5
P0001 1.0 ---
P0002 1.0 ---
mixed primers --- 1.0
2x GoTaq 12.5 12.5
dH2O 10.0 11.0
  25.0 25.0



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