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Catalog number: 170053, 170057

Components: 10x50 ml cells (170053), 20x50ml (170057), 10ml pUC18 (0.1ng/ml), 10ml SOC

Efficiency: ³5.0 x 10⁹ CPU (colony per mg pUC18)

Tube color: natural with yellow cap

Lot number: on box

Certified by:

Shelf life: 6 months from receipt date

Test conditions: transformations are performed with 50 ml aliquots of cells and 2 ml of pUC18 control plasmid (10 pg/ml) and without pUC18 following the protocol below with 1hour incubation at 37°C shaker. Dilute the reaction 1:200 with RT SOC and 100 ml of the dilution is plated in duplicate on LB agar plates with 50 mg/ml ampicillin. The plates are incubated at 37°C overnight and the efficiency is calculated based on the average number of colonies per plate.

Shipping conditions: on dry ice.

Storage conditions: immediately upon receiving at the bottom of a –80°C freezer. 

Transformation Protocol

1.        Thaw competent cells on ice. It is essential that cells be completely thawed before pipetting. Pipet slowly.

2.        Transfer 50 ml cells to a prechilled Falcon 352059 tube, add DNA of your choice (pure DNA or ligation mix) to the cells and incubate on ice for 30 minutes (may use 5 min for subcloning). Heat-shock at 42°C for 30 seconds and put the tube on ice for 2 minutes.

3.        Add 950 ml SOC and shake the cells at 37°C at 225 rpm for 30-60 minutes.

4.        Plate out desired amount on a prewarmed (37°C) plate with the appropriate antibiotic and incubate in a 37^oC incubator.

About ADD5a^TM cells

Our 5.0x10⁹ CPU ADD5a^TM competent cells, unlike other competent cells which claim similar CPU, carry no common antibiotics resistance and therefore can be used with E coli plasmid with any common antibiotics resistance. They are appropriate for cDNA library construction and routine subcloning. Cells provide a–complementation of the b-galactosidase gene for color selection and carry recA1 and endA1 makers, which minimizes recombination and enhance plasmid quality. Its genotype is endA1 recA1 relA1 gyrA96 hsdR17(r_k^-, m_k⁺) phoA supE44 thi-1 D(lacZYA-argF)U169 F80 D(lacZ)M15 F^-. Comparable to DH5a, a trademark of LTI.

5 min Transformation Protocol

1.    Thaw competent cells on ice.

2.    Transfer 50 ml cells to a prechilled microcentrifuge tube or Falcon 352059 tube, add DNA of your choice (pure DNA or ligation mix) to the cells and incubate on ice for 5 minutes.

3.    Plate out desired amount on a prewarmed (37°C) plate with the appropriate antibiotic and incubate in a 37°C incubator.

The transformation efficiency of the quick protocol is lower than that of the regular protocol but should be sufficient for subcloning.

Related Products:

Cat #: 30015M, Fastgrow^TM plate mix, for 500ml volume, sufficient to make 18-20 plates (100mm x 15mm). It can accelerate E. coli growth by 2-4 hr to pickable size on plate, depending on strains.

Cat #: 30025M, Fastgrow^TM liquid mix, for 500ml. It can accelerate E. coli growth in liquid.

Cat#: 170017, Fastgrow^TM 1x10⁹ chemically competent E. coli cells.

Cat#: 170047, ADD5a^TM ³1x10⁹ chemically competent E. coli cells.

Cat#: 170067, Bac-King^TM ≥1x10¹⁰ chemically competent E coli cells.

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Last modified: 04/25/06