IGEM:Harvard/2006/Cyanobacteria/Notebook/2006-8-20
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Miniprep of J04500
Played Jazz in the lab & miniprepped .2 mL J04500 as per the todo list left yesterday.
Notes: Eluted in H20
Left 4 miniprep tubes in our fridge. nanodrop results as follows: 1: 12.6 2: 13.2 3: 15.6 4: 14.2
The results are just about what we get every time.
Digests will follow later after dinner.
Anyone want to take a crack at the presentation for tomorrow while I do the digests? Or anyone want to do the digests and I'll take care of the presentation? Let me know.
Digestion of J04500 with X-P & S-P, KaiA/B/C with X-P
Performed the digests of the midiprepped KaiA/B/C and the miniprepped J04500 from earlier today. Ran two digests of each of the following: J04500 with X-P J04500 with S-P KaiA/B/C with X-P
J04500 Digest Protocol
Reaction X-P:
- 21.25 µL DNA
- 2.5 µL NEBuffer 3
- 0.5 µL XbaI
- 0.5 µL PstI
- 0.25 µL 100x BSA
Reaction S-P:
- 21.25 µL DNA
- 2.5 µL NEBuffer 2
- 0.5 µL SpeI
- 0.5 µL PstI
- 0.25 µL 100x BSA
Master X-P:
- 10 µL NEBuffer 3
- 2 µL XbaI
- 2 µL PstI
- 1 µL 100x BSA
Master S-P:
- 10 µL NEBuffer 2
- 2 µL SpeI
- 2 µL PstI
- 1 µL 100x BSA
Pipette 3.75 µL into each reaction. Digested for 16 hours at 37C in PCR-?.
KaiA/B/C X-P Digest Protocol
Reactions X-P:
- 8 µL midiprepped DNA
- 2.5 µL NEBuffer 3
- 0.5 µL XbaI
- 0.5 µL PstI
- 0.25 µL 100x BSA
- 13.25 µL H2O
Master X-P:
- 17.5 µL NEBuffer 3
- 3.5 µL XbaI
- 3.5 µL PstI
- 1.75 µL 100x BSA
- 92.75 µL H2O
Pipette 17 µL into each reaction. Digested for 16 hours at 37C in PCR-?.
Diagnostic digest of 13 minipreps from 2006-8-17 ligation
We performed \X-P digests of the 13 minipreps as per yesterday's to-do list.
Reaction:
- 21.25 µL DNA
- 2.5 µL NEBuffer 3
- 0.5 µL XbaI
- 0.5 µL PstI
- 0.25 µL 100x BSA
Master:
- 37.5 µL NEBuffer 3
- 7.5 µL XbaI
- 7.5 µL PstI
- 3.75 µL 100x BSA
Pipette 3.75 µL into each reaction.
Digested for 8 hours at 37C in PCR-1.
