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Team:Tsinghua/Notebook/24 September 2010
From 2010.igem.org
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== Module Ⅱ, group 4 == | == Module Ⅱ, group 4 == | ||
| + | Transform the trans5α with OmpA and CipC, respectively. | ||
| + | |||
| + | Transform the trans5α with pET21b. | ||
| + | |||
| + | Inoculation of OmpA、CipC、pET21b, 37℃，12h | ||
| + | |||
| + | Plasmid abstraction (OmpA、CipC、pET21b) | ||
| + | |||
| + | Measuring the concentration of the three kinds of plasmids. | ||
| + | |||
After getting the primer,Use the K-up and K-do to detect the P+K+C. Meanwhile use the digestion protocol to ensure the result. | After getting the primer,Use the K-up and K-do to detect the P+K+C. Meanwhile use the digestion protocol to ensure the result. | ||
Revision as of 02:39, 28 October 2010
Module Ⅱ, group 4
Transform the trans5α with OmpA and CipC, respectively.
Transform the trans5α with pET21b.
Inoculation of OmpA、CipC、pET21b, 37℃，12h
Plasmid abstraction (OmpA、CipC、pET21b)
Measuring the concentration of the three kinds of plasmids.
After getting the primer,Use the K-up and K-do to detect the P+K+C. Meanwhile use the digestion protocol to ensure the result.
Double digestion system:
P+E+M:
H2O 5.5-10.5μl buffer BamHI 2μl P+E+M 5-10μl SalI 1.5μl BamHI 1μl Total 20μl
P+K+C:
H2O 0-10.5μl buffer BamHI 2μl P+K+C 5-16μl KpnI 1.5μl BamHI 1μl Total 20μl
Run a gel. Order: P+E+M 1-6 double digestion, Marker, P+K+C 1-6 double digestion, P+K+C PCR.
result
P+E+M② is positive. It seems that PKC 1-6 are positive. But it still remains unclear whether the PCR bands are so weak and why the digestion result seemed negative.
