IGEM:Peking/2007/Count-Conjugation-Notebook/2007-8-19: Difference between revisions
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==colony PCR Test for R751, pSC101== | ==colony PCR Test for R751, pSC101== | ||
*Test Lb- R751 plate, Lb- pSC101 have the correct plasmid. | *Test Lb- R751 plate, Lb- pSC101 have the correct plasmid. | ||
*Test plate:LB- R751, Lb- pSB101, Tc+ R751-pSC101, Tc+ | *Test plate:LB- R751, Lb- pSB101, Tc+ R751-pSC101, Tc+ Dh5α-pSC101, Tc+ R751-pSC101, Amp+ Dh5α-R0040 | ||
*primer :R751 OriT primer, pSC101 primer. | |||
*according to <colony PCR STANDARD PROTOCOL> | |||
*PCR system contains(each well): | |||
<pre> | |||
0.5 µl Primer 1(100uM) | |||
0.5 µl Primer 2 | |||
2 µl dNTP(2.5uM) | |||
2.5 µl 10X Taq Buffer | |||
0.25 µl Taq | |||
19 µl dH20 | |||
1 µl template | |||
-------------------------- | |||
~25 µl Total | |||
</pre> | |||
Revision as of 23:18, 19 August 2007
Tandem Ori-T by Qu Mingzhi, Ren Ze
mini-prep R-OriT, S-Orit (pSB1A2)
- using Transgen mini plasmid puriflication kit.
- 50µL after purflication
mini-prep double digesting test
- R-OriT, S-OriT digestion system contains(Standard Assembly fragment):
4 µl 10*H buffer 1 µl EcoRI 1 µl PstI 20 µl Plasmid 14 µl dH20 -------------------------- 40 µl Total
electrophoresis result
colony PCR Test for R751, pSC101
- Test Lb- R751 plate, Lb- pSC101 have the correct plasmid.
- Test plate:LB- R751, Lb- pSB101, Tc+ R751-pSC101, Tc+ Dh5α-pSC101, Tc+ R751-pSC101, Amp+ Dh5α-R0040
- primer :R751 OriT primer, pSC101 primer.
- according to <colony PCR STANDARD PROTOCOL>
- PCR system contains(each well):
0.5 µl Primer 1(100uM) 0.5 µl Primer 2 2 µl dNTP(2.5uM) 2.5 µl 10X Taq Buffer 0.25 µl Taq 19 µl dH20 1 µl template -------------------------- ~25 µl Total
