Haynes Lab:Notebook/Investigating Photo-Switchable Synthetic Nucleosomes/2012/11/09

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(November 9, 2012)
Current revision (07:45, 14 November 2012) (view source)
(November 9, 2012)
 
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* 72°C, 3 min.
* 72°C, 3 min.
* 4°C ∞
* 4°C ∞
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{| class="wikitable" border=0
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| <u>Expected:</u><br>1. H2B = 410 <br>2. LOV = 461
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<center>
<center>
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[[Image:Gel_11-9-12.png|150px|Results of the digests are shown.]]
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[[Image:Gel_11-9-12.png|Results of the digests are shown.]]
[[Image:KAH_Fermentas_GeneRuler_1kbplus.jpg]]
[[Image:KAH_Fermentas_GeneRuler_1kbplus.jpg]]
</center>
</center>
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<center>
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Results of the PCR product digests are shown, with H2B on the left and LOV on the right. The gel was loaded with 5 μL samples.
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</center>
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November 9, 2012

  • First, resuspend dry primer oligos (in blue-capped tubes) to 100 μM
  • Then make working solutions of 10 μM
  • Reactions:
  1. H2B-GFP plasmid + BB_H2B fwd + BB_H2B rev
  2. LOV plasmid + BB_LOV fwd + BB_LOV rev
Reagents H2B LOV
Plasmid DNA 0.5 μL 0.5
primer 1 (10 μM) 1.0 1.0
primer 2 (10 μM) 1.0 1.0
2x GoTaq mix 12.5 12.5
dH2O 10.5 10.5

PCR program:

  • 95°C, 3 min.
  • [95°C, 30 sec; 57°C, 30 sec.; 72°C, 30 sec.] x35 cycles
  • 72°C, 3 min.
  • 4°C ∞
Expected:
1. H2B = 410
2. LOV = 461

Results of the digests are shown. Image:KAH_Fermentas_GeneRuler_1kbplus.jpg

Results of the PCR product digests are shown, with H2B on the left and LOV on the right. The gel was loaded with 5 μL samples.



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