Super-Comp
Media and Reagents for Preparing Super-Competent Host Cells

BIO 101 introduces a procedure and reagents for preparing highly competent host cells without the necessity of involved procedures, such as the effective but intensive methods described by Hanahan [J. Mol. Biol.(1983) 166, 557]. An important step in the BIO 101 protocol is to grow the cells in SUPER-COMP media. SUPER-COMP is an extremely rich medium that significantly increases the competency of the cells when using a simple RbCa means of preparing competent host cells.

Protocol for preparing Super-Comp Cells.

1. Dilute 1 ml of a fresh overnight culture of host cells grown in LB medium into 25 ml of 37°C SUPER-COMP Broth media in a 500 ml flask.
2. Incubate cells with vigorous aeration for 1-2 hours until cells reach mid-log phase (approximately 8 x 10⁸ cells/ml, OD600 reading 0.8 for most E. coli host cells).
3. Centrifuge culture in a sterile tube at 4°C to pellet cells. Resuspend pellet in 15 ml of ice cold 1 x RbCa TXN SALTS.
4. After 15 minutes on ice, centrifuge to pellet cells and gently resuspend pellet in 2 ml of ice cold 1 x RbCa TXN SALTS. Keep cell suspension on ice.

The SuperComp cells are immediately ready for DNA uptake, or for maximum transformation efficiency, store for 12-24 hours at 4°C to increase competency 4-6 fold (at 48 hours they will be approximately as efficient as when first made). Competent cells can also be frozen in aliquots at -80°C after adding glycerol to 20%.