Intact Genomics EHA105 ElectroComp Agrobacterium cells are optimized for the highest transformation efficiencies which is ideal for applications requiring high transformation efficiencies, such as with cDNA or gDNA library construction. The EHA105 strain is useful for transgenic operations of rice, tobacco and other plants. EHA105 contains a rifampicin resistance gene (rif). EHA105 strain also contains an amber basic Ti plasmid pEHA105 (pTiBo542DT-DNA) without self-transport function, which contains the vir gene.
SpecificationsCompetent cell type: ElectrocompetentSpecies: A. tumefaciensStrain: EHA105Format: TubesTransformation efficiency: ≥ 1 x 107 cfu/µg pCAMBIA1391z DNABlue/white screening: NoShipping condition: Dry ice
Reagents Needed for One ReactionEHA105 ElectroComp Agrobacterium: 25 µlDNA (pCAMBIA1391z, 500 pg/µl): 1 µlRecovery medium: 1 ml
StorageEHA105 ElectroComp Agrobacterium: -80 ºCpCAMBIA1391z control DNA: -20 ºCRecovery medium: 4 ºC
Quality ControlTransformation efficiency is tested by using the pCAMBIA1391z control DNA supplied with the kit and using the protocol in this manual. Transformation efficiency should be ≥1 x 107 CFU/µg pCAMBIA1391z DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.
General GuidelinesFollow these guidelines when using EHA105 ElectroComp Agrobacterium:
- 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 such as Bio-Rad Gene Pulser II #165-2105, capable of generating field strengths of 16 kV/cm is required.
Calculation of Transformation EfficiencyTransformation Efficiency (TE) is defined as the number of colony forming units (cfu) produced by transforming 1µg of plasmid into a given volume of competent cells.TE = Colonies/µg/PlatedTransform 1 µl of (500 pg/µl) pCAMBIA1391z control plasmid into 25 µl of cells, add 974 µl of Recovery Medium. Recover for 3 hours and plate 100 µl. Count the colonies on the plate in two days. If you count 500 colonies, the TE is calculated as follows:Colonies = 500µg of DNA = 0.0005Dilution = 100/1000 = 0.1TE = 500/.0005/.1 = 1×1071284-12 1
Please note, all agrobacterial strains are not well studied for antibiotic resistance and there are many agrobacterial strains. Therefore, it is the customer’s responsibility to make sure his/her vectors are compatible with the Agrobacterial strains if he/she uses an alternate antibiotic selection than kanamycin-selection.1284-12 1284-36 1284-20
