User:Karmella Haynes/Notebook/BioBrick cloning/2015/05/20: Difference between revisions

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| bgcolor=#cfcfcf | Vol
| bgcolor=#cfcfcf | Vol
| bgcolor=#cfcfcf | Mix (x6)
| bgcolor=#cfcfcf | Mix (x6)
| rowspan=7 | Expected:<br>1. pAubR = 153<br>2. pBjaR = 122<br>3. pBraR = 214<br>4. pRpaR = 136
| rowspan=7 | Expected:<br>1. H3-1-40 = 129<br>2. H3-1-60 = 189<br>3. H3-1-80 = 250<br>4. H3-1-116 = 373<br>5. H3-1-136 = 433
| rowspan=7 | [[Image:KAH050515_gel2.jpg|250px|Hover name]]<br>5 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| rowspan=7 | [[Image:KAH052015_gel1.jpg|250px|Hover name]]<br>5 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
|-
|-
| gBlock DNA (2 ng/μL) || 0.5  || 3.0  
| gBlock DNA (2 ng/μL) || 0.5  || 3.0  

Revision as of 16:48, 21 May 2015

Karmella's BioBrick Cloning <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
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05/20/15

  • Cas-tone - Phusion PCR for H3


Cas-tone - Phusion PCR for H3

  • Phusion PCR-amplify promoters
  1. mEmerald-H3-23, H3_1 f1 / H3_40 r1
  2. mEmerald-H3-23, H3_1 f1 / H3_60 r1
  3. mEmerald-H3-23, H3_1 f1 / H3_80 r1
  4. mEmerald-H3-23, H3_1 f1 / H3_116 r1
  5. mEmerald-H3-23, H3_1 f1 / H3_136 r1
Reagent Vol Mix (x6) Expected:
1. H3-1-40 = 129
2. H3-1-60 = 189
3. H3-1-80 = 250
4. H3-1-116 = 373
5. H3-1-136 = 433 		Hover name
5 μL/lane; 1% agarose; Ladder
gBlock DNA (2 ng/μL) 0.5 3.0
10 μM H3_1 f1 1.0 6.0
10 μM r primer 1.0 ---
10 mM dNTPs 1.0 6.0
5x HF buffer 10.0 60.0
Phusion Pol. 0.5 3.0
dH2O 36.0 216.0
  50.0 μL

Thermal cycler: Bio-Rad - Phusion

  • 98°C 3 min.
  • 30x[98°C, 10 sec; 70°C 30 sec; 72°C 30 sec]
  • 72°C 3 min.
  • 4°C, ∞





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