JM109 Chemically Competent E. coli Cells
GoldBio’s JM109 Chemically Competent E. coli cells are suitable for high efficiency transformation in a wide variety of routine applications such as plasmid isolation, library construction, cloning, and subcloning. JM109 is a K strain bacterium that carries multiple features including the ɸ80lacZ∆M15 marker, which provides α-complementation of the β-galactosidase gene with blue/white screening protocol as well as the hsdR17 genotype which prevents endogenous endonucleases from cleaving DNA.
Competent cell type: Chemically competent
Derivative of: JM109
Species: E. coli
Transformation efficiency: ≥6 x 105 cfu/µg pUC19 DNA
Blue/white screening: Yes
Storage/Handling: This product may be shipped on dry ice. JM109 Chemically Competent E. coli cells should be stored at -80°C, pUC19 Control DNA should be stored at -20°C and recovery medium should be stored at 4°C immediately upon arrival. When stored under the recommended conditions and handled correctly, these products should be stable for at least 1 year from the date of receipt.Genotype
endA1, recA1, gyrA96, thi, hsdR17 (rk–, mk+), relA1, supE44, Δ( lac-proAB), [F´ traD36, proAB, lacIqZΔM15]
Reagents Needed for One Reaction
- JM109 chemically competent cells: 50 µl
- DNA (or pUC19 Control, 10 pg/µl): 1 µl
- Recovery medium: 1 ml
Transformation efficiency is tested by using the pUC19 control DNA supplied with the kit and using the protocol given below. Transformation efficiency should be ≥6 x 10 5 CFU/µg pUC19 DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.
- Handle competent cells gently as they are highly sensitive to changes in temperature or mechanical lysis caused by pipetting.
- Thaw competent cells on ice, and transform cells immediately following thawing. After adding DNA, mix by tapping the tube gently. Do not mix cells by pipetting or vortexing.
Calculation of Transformation Efficiency
Transformation Efficiency (TE) is defined as the number of colony forming units (cfu) produced by transforming 1 µg of plasmid into a given volume of competent cells.
- TE = Colonies/µg/Dilution
- Colonies = the number of colonies counted
- µg = amount of DNA transformed in µg
- Dilution = total dilution of the DNA before plating
Example: Transform 1 µl of (10 pg/µl) control plasmid into 25 µl of cells, add 975 µl of Recovery Medium. Dilute 10 µl of this in 990 µl of Recovery Medium and plate 50 µl. Count the colonies on the plate the next day. If you count 250 colonies, the TE is calculated as follows:
Colonies = 250
µg of DNA = 0.00001
Dilution = 10/1000 x 50/1000 = 0.0005
TE = 250/0.00001/0.0005 = 5.0 × 1010
|Storage/Handling||Store Competent Cells at -80°C.|