IGEM:Harvard/2006/Cyanobacteria/Notebook/2006-8-2: Difference between revisions
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==X-S digest of KaiA/B and RFP BioBrick plasmid== | ==X-S digest of KaiA/B and RFP BioBrick plasmid== | ||
We performed X-S digests of KaiA/B and the RFP BioBrick plasmids. | We performed X-S digests of KaiA/B and the RFP BioBrick plasmids. There were 6 digests in total: LC3A KaiA, LC3A KaiB, LC3B KaiA, LC3B KaiB, RFP #3, and RFP #4 (RFP numbers refer to the 4 minipreps we did yesterday). | ||
Reaction: | Reaction: | ||
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==Gel images from yesterday's AU PCR== | ==Gel images from yesterday's AU PCR== | ||
[[Image:2006-8-2 entire sequence PCR.jpg|thumb|left|Click for legend]] | [[Image:2006-8-2 entire sequence PCR.jpg|thumb|left|Click for legend]] | ||
==Phosphotase treatment of KaiA/B and RFP BioBrick plasmid digestions== | |||
After completing the X-S digests, we added 1 µL alkaline phosphotase to each digestion and incubated for 1 hour at 37C. |
Revision as of 14:12, 2 August 2006
X-S digest of KaiA/B and RFP BioBrick plasmid
We performed X-S digests of KaiA/B and the RFP BioBrick plasmids. There were 6 digests in total: LC3A KaiA, LC3A KaiB, LC3B KaiA, LC3B KaiB, RFP #3, and RFP #4 (RFP numbers refer to the 4 minipreps we did yesterday).
Reaction:
- 20 µL DNA
- 2.5 µL Buffer 2 (from NEB)
- 0.25 µL BSA (100x)
- 0.5 µL XbaI enzyme
- 0.5 µl SpeI enzyme
- 1.25 µL H2O
Incubate for 2 hours at 37C, heat inactivate for 20 minutes at 80C, hold at 4C.
Gel images from yesterday's mutagenic PCR
Gel images from yesterday's AU PCR
Phosphotase treatment of KaiA/B and RFP BioBrick plasmid digestions
After completing the X-S digests, we added 1 µL alkaline phosphotase to each digestion and incubated for 1 hour at 37C.