Haynes Lab:Notebook/Engineering PC-TFs/2013/01/25

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| bgcolor=#cfcfcf | Volume  
| bgcolor=#cfcfcf | Volume  
| rowspan="7" | <u>Expected:</u><br>1. pSB1A3 = ~2000<br>2. hPCD = 186<br>3. BL01 = 2520
| rowspan="7" | <u>Expected:</u><br>1. pSB1A3 = ~2000<br>2. hPCD = 186<br>3. BL01 = 2520
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| rowspan="7" | [[Image:image.jpg|200px|PCR results]]<br>10 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
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| rowspan="7" | [[Image:gg_gel1-25-2013|200px|PCR results]]<br>10 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
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|-
| DNA(plasmid) || 0.2 μL
| DNA(plasmid) || 0.2 μL

Revision as of 01:25, 28 January 2013

Engineering PC-TFs Main project page
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01/25/13

  • Type IIS assembly - day 1


PCR

  • Resuspend new primers to a final concentration of 100 μM
  • Make 10 μM dilutions in separate tubes
  • All of the reactions are as follows:
  1. pSB1A3, gg0001, gg0002
  2. template, primer 1, primer 2
  3. template, primer 1, primer 2
  4. template, primer 1, primer 2


Reagent Volume Expected:
1. pSB1A3 = ~2000
2. hPCD = 186
3. BL01 = 2520 		Image:Gg gel1-25-2013
10 μL/lane; 1% agarose; Ladder
DNA(plasmid) 0.2 μL
primer 1 1.0
primer 2 1.0
2x GoTaq 25.0
dH2O 22.8
  50.0 μL
  • 95°C, 3 min.
  • [95°C, 30 sec.; 57°C, 30 sec.; 72°C, 1 min.] x35
  • 72°C, 3 min.
  • 4°C, ∞


Purification

  • Clean with Zymo DNA c&c; elute w/ 20 μL dH2O
  • Measure [DNA]
Sample OD 260 260/280 ng/μL
1. pSB1A3 ### ### ###
2. ### ### ### ###
3. ### ### ### ###



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