IGEM:MIT/2007/Notebook/2007-7-17: Difference between revisions
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==Digestion of CPX for 3A== | ==Digestion of CPX for 3A== | ||
<pre> | <pre> | ||
5µl CPX (diluted to 1µg/5µl) | |||
0.5µl BSA | 0.5µl BSA | ||
5µl NEB3 Buffer | 5µl NEB3 Buffer | ||
1µl XbaI | 1µl XbaI | ||
1µl PstI | 1µl PstI | ||
37.5µl H2O | |||
------------------ | ------------------ | ||
50µl Total | 50µl Total | ||
</pre> | </pre> | ||
==Digestion of (F2620 and B0034) for 3A== | ==Digestion of (F2620 and B0034) for 3A== | ||
Revision as of 07:21, 17 July 2007
Agenda
- Check sequencing results of second (uncontaminated) set of miniprepped 3K3+F2620+B0034
- Digest CPX, 3K3+F2620+B0034 (if sequencing is correct), and a hi-copy plasmid for 3A assembly
- Ligate digested CPX, 3K3+F2620+B0034, and hi-copy plasmid
- Test Standard Assembly of pSB3k3 with F2620, B0034, E1010 using AHL
- LC grown overnight
Digestion of CPX for 3A
5µl CPX (diluted to 1µg/5µl) 0.5µl BSA 5µl NEB3 Buffer 1µl XbaI 1µl PstI 37.5µl H2O ------------------ 50µl Total
Digestion of (F2620 and B0034) for 3A
20µl (F2620 and B0034) @ 35ng/µl 0.5µl BSA 5µl NEB2 Buffer 1µl EcoRI 1µl SpeI 22.5µl H2O ------------------ 50µl Total
- Digested pSB1AC3: 15µl @ 125 ng/µl
