Difference between revisions of "Team:UCLA/Notebook/Honeybee Silk/11 August 2015"
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+ | ='''Ligation of psb1c3+T7 promoter and Silk+GFP'''= | ||
+ | |||
+ | {| class="wikitable" style="text-align:center; width:400px; height:200px;" | ||
+ | |+ | ||
+ | |- | ||
+ | ! Component | ||
+ | ! Volume (out of 20uL) | ||
+ | |- | ||
+ | ! T4 Ligase Reaction Buffer | ||
+ | | 2 ul | ||
+ | |- | ||
+ | ! Backbone (psb1c3+t7 promoter) | ||
+ | | 0.8uL (50 ng) | ||
+ | |- | ||
+ | ! Insert (silk + spyC) | ||
+ | | 1.5 ul (146 ng) | ||
+ | |- | ||
+ | ! H20 | ||
+ | | 14.7 ul | ||
+ | |- | ||
+ | ! Ligase | ||
+ | | 1uL | ||
+ | |} | ||
+ | *Program =16C overnight, 10 minutes heat inactivation at 65C. | ||
+ | |||
=8/11= | =8/11= | ||
===Protein Yield=== | ===Protein Yield=== |
Latest revision as of 22:02, 24 August 2015
Ligation of psb1c3+T7 promoter and Silk+GFP
Component | Volume (out of 20uL) |
---|---|
T4 Ligase Reaction Buffer | 2 ul |
Backbone (psb1c3+t7 promoter) | 0.8uL (50 ng) |
Insert (silk + spyC) | 1.5 ul (146 ng) |
H20 | 14.7 ul |
Ligase | 1uL |
- Program =16C overnight, 10 minutes heat inactivation at 65C.
8/11
Protein Yield
- Yield from 8/7 silk purification
- BCA assay
- Based on the interpolated Absorbencies from the BCA assay the concentration of our 8/7 silk is approximately 0.78 mg/ml in around 12 ml of solution, giving a conservative 9 total mg of protein for a 300 ml culture.
- This is an improvement over the 7/29 purification where we got around 1.6 mg/ml in only 0.75 ml of solution from a 100 ml culture.