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Catalog number: 170043, 170045, 170049

Components: 10x50ul cells (170043), 5x0.2ml cells (170045), 20x50ul cells (170049), 10ul pUC18 (0.1ng/ul), 10ml SOC

Efficiency: ³1.0 x 10⁹ CPU (colony per ug 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 100 ul aliquots of cells and 2 ul of pUC18 control plasmid (10 pg/ul) and without pUC18 following the protocol below with 1 hour incubation at 37°C shaker. Dilute the reaction 1:100 with SOC and 100 ul of the dilution is plated in duplicate on LB agar plates with 50 ug/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. t is essential that cells be completely thawed before pipetting. Pipet slowly.

2.                  Aliquot 100 ul 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 45 seconds and put the tube on ice for 1.5-2 minutes.

3.                  Add 900 ul SOC or 950 ul SOC if using 50 ul cells 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  cells

ADD5a  competent cells 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. Aliquot 100 ul 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#: 170015, Fastgrow^TM ³1x10⁹ chemically competent E. coli cells.

Cat#: 170055, ADD5a^TM ³5x10⁹ chemically competent E. coli cells.

Cat#: 170067, Bac-King^TM ³1x10¹⁰ chemically competent E. coli cells.

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Last modified: 04/25/06