Team:UCLA/Notebook/Recombinant Expression
Recombinant Silk Functionalization
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Today we began cloning our GFP.
- PCR'd off template
- Ran gel
- Restriction digest
- Ligated into backbone
For our ligation, we made the following modifications:
- Tried it with newly bought ligase
- Left reaction overnight instead of 2 hrs
- Vector to insert ratio was 1:5 instead of 1:3
PCR Reaction:
| Component | Volume |
| 5X Q5 Reaction Buffer | 5 |
| 10 mM dNTPs | 0.5 |
| 10 uM Forward (primer 3/7) | 1.25 |
| 10 uM Reverse (primer 8) | 1.25 |
| Template (diluted to 1ng/uL) | 0.5 |
| Q5 High Fidelity DNA Polymerase | 0.25 |
| Nuclease Free Water | 16.25 |
Gel: Lot of bands, all at correct sizes
Contents
Goals
Goals
- Finish Purification of ABD containing construct
- Create hydrogel with ABD and B.mori silk and see if it retains/binds albumin
Achievements
Achievements
- Cloned the ABD construct and begun purification methods
What to accomplish next
Julian
- Test albumin binding properties of the ABD silk construct
- test hydrogel formation and albumin retention
Raw lab notebook entries
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