DH5-alpha Electrocompetent E. coli Cells
GoldBio’s DH5-alpha Electrocompetent E. coli cells are suitable for high efficiency transformation in a wide variety of routine applications such as plasmid isolation, cloning, and subcloning. Mutations in endA1 and recA1 ensure increased plasmid yield and improved plasmid quality.
Competent cell type: ElectroCompetent
Derivative of: DH5-alpha
Species: E. coli
Transformation efficiency: ≥2 x 1010 cfu/µg pUC19 DNA
Blue/white screening: Yes
Storage/Handling: This product may be shipped on dry ice. DH5-alpha Electrocompetent 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.
- ≥2 x 1010 cfu/µg efficiency with electroporation
- Greatly increased plasmid yield and quality due to endA1 mutation
- High-efficiency transformation with plasmids 30 kb in size
- Blue/white screening of recombinant clones due to lacZΔM15
- Ensured insert stability due to recA1 mutation
Φ80 Δ(lacZ)M15 fhuA2 Δ(argF-lacZ)U169 phoA glnV44 gyrA96 recA1 relA1 endA1 thi-1hsdR17
Reagents Needed for One Reaction
- DH5-alpha electrocompetent cells: 25 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 ≥2 x 1010 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.
Note: A high-voltage electroporation apparatus capable of generating field strengths of 16 kV/cm is required.
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
Procedure for transformation through electroporation utilizing DH5-alpha Electrocompetent E. coli Cells.
Guide to GoldBio's competent cell collection containing transformation efficiencies, characteristics and applications for each of our competent cell lines.
Chart containing genotypes and transformation efficiencies for GoldBio's competent cell collection.
This guide provides nomenclature information relating to genotypes and genetic markers of E. coli and competent cells.