Team:Calgary/2 July 2009
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+ | *Protocol: Used GenElute Plasmid Miniprep Kit (Sigma), lot # 048k6062, used as according to the manufacturers instructions, eluting in 50 μL ddH2O. | ||
+ | |||
+ | *Results: | ||
+ | |||
+ | {| border="1" | ||
+ | | '''Plasmid''' || '''260/280''' || '''260/230''' || '''Concentration [ng/μL]''' | ||
+ | |- | ||
+ | | ''psB1AC3 1 || 1.96 || 3.48 || 76.9 | ||
+ | |- | ||
+ | | ''psB1AC3 2 || 1.93 || 2.29 || 98.1 | ||
+ | |- | ||
+ | | ''psB1AC3 3 || 1.87|| 1.88|| 48.5 | ||
+ | |- | ||
+ | | ''psB1AC3 4|| 1.90|| 2.11|| 118.9 | ||
+ | |- | ||
+ | | ''psB1AK3 1 || 2.05 || 2.42 || 134.4 | ||
+ | |- | ||
+ | | ''psB1AK3 2 || 1.95 || 2.23 || 55.6 | ||
+ | |- | ||
+ | | ''psB1AK3 3 || 1.91 || 2.22 || 97.9 | ||
+ | |- | ||
+ | | ''psB1AK3 4 || 2.03 || 1.92 || 44.5 | ||
+ | |} | ||
+ | |||
+ | Verification digest to verify the presence of restriction sites in the psB1AC3, psB1AK3 vectors as well as in our TOPO TA vector with LuxOD47E. | ||
+ | |||
+ | *Protocol: | ||
+ | <Br> | ||
+ | 5 reactions for psB1AC3 vector, 5 reactions for psB1AK3 vector, 1 reaction for gene of interest (LuxOD47E in pCR2.1-TOPO vector). | ||
+ | <Br> | ||
+ | BBK Vector Reaction List | ||
+ | <Br> | ||
+ | Reaction 1- Not1 + React 3 Buffer | ||
+ | Reaction 2- EcoRI + SpeI + React Buffer 4 | ||
+ | Reaction 3- XbaI + PstI + React 2 Buffer | ||
+ | Reaction 4- EcoRI + PstI + React 2 Buffer | ||
+ | Reaction 5- XbaI + SpeI + React 4 Buffer | ||
+ | <Br> | ||
+ | BBK vector tube preparation | ||
+ | 8 uL Plasmid (psB1AC3 or psB1AK3) | ||
+ | 2 uL appropriate React Buffer | ||
+ | 1 uL of each appropriate restrcition enzyme | ||
+ | ddH20 up to 20 uL | ||
+ | <Br> | ||
+ | TOPO vector tube preparation | ||
+ | 10 uL DNA (LuxOD47E in pCR2.1- TOPO vector) | ||
+ | 7 uL ddH20 | ||
+ | 2 uL React 3 Buffer | ||
+ | 1 uL NotI restriction enzymes | ||
+ | <Br> | ||
+ | Left to digest overnight in waterbath at 37°C. | ||
+ | |||
+ | <Br> | ||
<html> | <html> | ||
+ | <Br> | ||
</div> | </div> | ||
</td> | </td> |
Revision as of 19:32, 29 July 2009
UNIVERSITY OF CALGARY