HB101 Chemically Competent E. coli Cells

Product Description

GoldBio’s HB101 chemically competent E. coli cells allow you to obtain high transformation efficiency in applications such cloning and sub-cloning. Our E. coli HB101 proceeds from the K12 x B hybrid strain. HB101 strain also contains the recA13 mutation useful in the insert stability and minimization of recombination. E. coli HB101 also has the hsdS20(rB-mB-) genotype that prevents the cleavage of cloned DNA by endogenous restriction enzymes.

Kit Components

  • Competent Cells
  • 1 x 12 mL Recovery Media
  • 1 x 25 µl Control Plasmid (pCAMBIA1391z, 10 ng/µl)

Product Specifications
Competent cell type: Chemically competent
Derivative of: HB101
Species: E. coli
Transformation efficiency: ≥7 x 106 cfu/µg pUC19 DNA
Blue/white screening: No

Storage/Handling: This product may be shipped on dry ice. HB101 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

F- Lambda- araC14 leuB6(Am) DE(gpt-proA)62 lacY1 glnX44(AS) galK2(Oc) recA13 rpsL20(strR) xylA5 mtl-1 thiE1 hsdS20(rB-, mB-)

Reagents Needed for One Reaction

  • HB101 chemically competent cells: 50 µl
  • DNA (or pUC19 Control, 10 pg/µl): 1 µl
  • Recovery medium: 1 ml

Quality Control
Transformation efficiency is tested by using the pUC19 control DNA supplied with the kit and using the protocol given below. Transformation efficiency should be ≥7 x 106 CFU/µg pUC19 DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.

General Guidelines

  • 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

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