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From 2010.igem.org

Module I, group 2(b)

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.