User:Robwarden/Notebook/Caspase-3 Activatable Probe/2009/09/10

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 * style="background-color: #EEE"|[[Image:Penn_fulllogo.gif|128px]] Caspase-3 Activatable Probe
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 * style="background-color: #F2F2F2" align="center"|  |Main project page


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Miniprep
with Qiagen kit

Digestion

 * 20 μL reaction volume
 * NEB Buffer 4
 * .5 μL of EcoRI
 * .5 μL of SgrAI
 * 10 μL of DNA
 * Incubate at 37°C for 1 hr

Agarose Gel

 * 1% Agarose, run for 30 min at 120V


 * Predicted Banding Patterns
 * Correct: 6535
 * Incorrect: 6297, 1434


 * All colonies appeared to work, except for #12

Sequencing
Colonies 2, 5, 7, and 13 were submitted for sequencing. T7 was used for the forward primer. BHG was used for the reverse primer. Both primers were supplied by the sequencing facility.

Reaction Setup
5.0 μL 10X Buffer 1.0 μL hRLuc8 plasmid 2.5 μL Primer "hRLuc8_EcoRI_Fwd" 2.5 μL Primer "hRLuc8_XhoI_Rev" 1.0 μL 10mM dNTP mix 0.5 μL Vent Polymerase 37.5 μL dH2O

Thermo Cycle

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