Haynes Lab:Notebook/Protein-DNA fusions in living microbes/2013/10/09: Difference between revisions
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used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf | used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf | ||
Made SOC media: | |||
LB miler broth + tryptone + 20 mM MgSO4 and 2.5 mM KCl (http://openwetware.org/wiki/SOB) | |||
Grew up 4 colonies of BL21(DE3) in Kan media, used 3 of them (1, 2, 4) to induce expression of Topo per protocol: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf | |||
Ran PCR using 1-omega F TSE and alpha-4 R SP and PSV plasmid template. PCR product has 3 topo cut sites. | |||
Ran gel of wheat-germ topo +RFP pSB1C3 plasmid. From left to right: 2uL, 1.5uL, 1uL, 0.5uL, blank glycerol stock (15/30 uL in each well) | |||
Made competent cell stock of BL21(DE3) from cells given by Kylie | |||
Used Bug buster on induced samples (1, 2, 4) per protocol: http://www.ebiotrade.com/buyf/productsf/novagen/70584-000.pdf | |||
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Revision as of 23:58, 2 December 2013
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Entry titlePerformed successful tranformation with Kylie's BL21(DE3) used protocol here: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf Made SOC media: LB miler broth + tryptone + 20 mM MgSO4 and 2.5 mM KCl (http://openwetware.org/wiki/SOB) Grew up 4 colonies of BL21(DE3) in Kan media, used 3 of them (1, 2, 4) to induce expression of Topo per protocol: http://www.molecularinfo.com/MTM/A/A3/A3-1.pdf Ran PCR using 1-omega F TSE and alpha-4 R SP and PSV plasmid template. PCR product has 3 topo cut sites. Ran gel of wheat-germ topo +RFP pSB1C3 plasmid. From left to right: 2uL, 1.5uL, 1uL, 0.5uL, blank glycerol stock (15/30 uL in each well) Made competent cell stock of BL21(DE3) from cells given by Kylie Used Bug buster on induced samples (1, 2, 4) per protocol: http://www.ebiotrade.com/buyf/productsf/novagen/70584-000.pdf | |
