Title: Transforming Bacteria with the LIC Reaction
Course: BTEC-1100-301-302-303-F21 > Section: Module 4 - Applied Molecular Biology

We will be using prepared competent cells for transformation.  Competent cells are bacterial cells that have been chemically treated so that they are capable of intaking the recombinant DNA. Cells that have received the pLATE31 plasmid are then selected for using the antibiotic ampicillin in the growth media.  Because the pLATE31 plasmid contains a gene that encodes a protein that breaks down ampicillin, only those bacteria harboring our plasmid will survive.  Your instructor will provide you with two 50 μl aliquots of Novablue competent cells for transformation of the LIC reaction and control reaction.

Transformation

1. Thaw cells on ice and mix gently to ensure that the cells are evenly suspended.
2. Add 5 μl of the LIC reaction directly to the cells. Stir gently to mix.
3. In a separate tube of Novablue cells add 5 μl of the control reaction and stir gently to mix.
4. Place the tubes on ice for 5 min.
5. Heat the tubes for exactly 30 sec in a 42°C heat block; do not shake.
6. Place on ice for 2 min.
7. Plate cells on LB/agar plates containing 100 μg/ml ampicillin and incubate overnight at 37°C.

Learning Objectives:

- Transform bacteria using the results of LIC.