DH10B BAC Library Electrocompetent E. coli Cells
GoldBio’s DH10B BAC Library Electrocompetent E. coli cells are suitable for demanding cloning applications ̶ BAC library construction and large construct transformation or cloning difficult targets ̶ requiring the greatest number of transformants possible.
Competent cell type: ElectroCompetent
Derivative of: DH10B™
Species: E. coli
Transformation efficiency: ≥5 x 1010 cfu/µg pUC19 DNA
Blue/white screening: Yes
Storage/Handling: This product may be shipped on dry ice. DH10B BAC Library 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.
- ≥5 x 1010 cfu/µg efficiency with electroporation.
F – mcrA ∆(mrr-hsdRMS-mcrBC) endA1 recA1 φ80dlacZ∆M15 ∆lacX74 araD139 ∆(ara, leu)7697 galU galK rpsL (StrR) nupG λ-
Reagents Needed for One Reaction
- DH10B BAC electrocompetent cells: 20 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 ≥5 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 DH10B BAC 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.