User:Mary Mendoza/Notebook/CHEM 571 Experimental Biological Chemistry I/2012/10/24

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Transformation of BL21 cells

The following procedures were conducted near the proximity of the Bunsen burner in accordance with aseptic techniques. The procedure is based from the laboratory Transformation protocol.

  • Transferred 5 μL of PCR plasmid in two sterilized tubes. One tube will be filled with LB media, the other with SOC media. Both media are from Novagen.
  • Added 40 μL of thawed BL21 cells into each tube. The tubes were immediately placed on a VWR analog heatblock at 42°C for 30 sec.
  • After the 30 sec. mark, the tubes were submerged in ice. This rapid change in temperatures is a method of heat shock. This is an adequate of method of allowing the cells to incorporate the mutated plasmid into their intracellular matrix.
  • Delivered 250 μL of the LB and SOC media using an automatic delivery pipet into the labeled tubes.
  • The tubes containing the media were centrifuge for 1 h. at 37°C by Dr. Miller.
  • During the latter of the lab period, 2 plates were inoculated for each cell grown in LB and SOC media.
  • The first plate was inoculated with 50 μL of the BL21 cell grown in LB; the second plate was inoculate with a greater volume of cells with 200 μL.
  • Dhea Patel plated the BL21 cells grown in SOC media.