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To prevent recircularization of digested plasmids, we used NEB's Antarctic Phosphatase according to protocol.<br> | To prevent recircularization of digested plasmids, we used NEB's Antarctic Phosphatase according to protocol.<br> | ||
<b>Gel Electrophoresis</b><br> | <b>Gel Electrophoresis</b><br> | ||
| - | To view lengths of DNA, Orange G 6x from NEB was added 1:3 to 300ng miniprepped DNA and run against 5uL of Hyperladder I from NEB in the lanes of 1% UltraPure agarose (Invitrogen) gels in TAE buffer at 120V for 45 minutes.<br> | + | To view lengths of DNA, Orange G 6x from NEB was added 1:3 to 300ng miniprepped (and sometimes digested) DNA and run against 5uL of Hyperladder I from NEB in the lanes of 1% UltraPure agarose (Invitrogen) gels in TAE buffer at 120V for 45 minutes.<br> |
<b>DNA Gel Extraction</b><br> | <b>DNA Gel Extraction</b><br> | ||
To extract the DNA from the agarose gel, the QIAquick Gel Extraction kit was used according to protocol.</td> | To extract the DNA from the agarose gel, the QIAquick Gel Extraction kit was used according to protocol.</td> | ||
Revision as of 16:06, 10 October 2010
iGEM 2010
Undergraduates Instructors Advisors Fun Sponsors
Overview Toggle Phage Mammalian Summary Acknowledgements
Materials & Methods Biosafety Journal Club
Bacterial Construction Protocol
Bacterial Experimental Protocol
Phage western blot
Mammalian Cell Lines
Microfluidic stress
Materials
biobrick construction |
Miniprep To extract DNA from cells, we used the QIAprep Spin Miniprep Kit according to protocol. DNA Digestion For both restriction mapping and for digesting a plasmid to insert or extract a biobrick, the appropriate buffer was added 1:10 to a DNA sample and 10 units of the appropriate enzyme(s) were used per 1ug of DNA. The reactions were allowed to run at the temperature optimal for the enzyme for 1 hour, the enzymes were denatured at 65°C for 20 minutes, and then the reactions were held at 16°C. Antarctic Phosphatase To prevent recircularization of digested plasmids, we used NEB's Antarctic Phosphatase according to protocol. Gel Electrophoresis To view lengths of DNA, Orange G 6x from NEB was added 1:3 to 300ng miniprepped (and sometimes digested) DNA and run against 5uL of Hyperladder I from NEB in the lanes of 1% UltraPure agarose (Invitrogen) gels in TAE buffer at 120V for 45 minutes. DNA Gel Extraction To extract the DNA from the agarose gel, the QIAquick Gel Extraction kit was used according to protocol. |
