Intact Genomics (ig®) MSU440 Electrocompetent Agrobacterium cells are made from a specific strain of Rhizobium rhizogenes (formerly Agrobacterium rhizogenes), Agrobacterium rhizogenes(str R) MSU440 Ri (agropine type). Agrobacterium rhizogenes is a soil-borne gram-negative bacterium that can infect most dicotyledons, a few monocotyledons and some gymnosperms. MSU440 Electrocompetent Agrobacterium are optimized for the highest transformation efficiencies and are useful for transgenic operations of corn, tobacco, wormwood, tea tree and other plants. MSU440 Agrobacterium rhizogenes strain contains agrobacterium-type Ri plasmid and displays streptomycin resistance.
SpecificationsCompetent cell type: ElectrocompetentSpecies: R. rhizogenesStrain: MSU440Format: TubesTransformation efficiency: ≥ 1 x 107 cfu/µg pCAMBIA1391z DNABlue/white screening: NoShipping condition: Dry ice
Reagents Needed for One ReactionMSU440 ElectroCompetent Agrobacterium: 25 µlDNA (pCAMBIA1391z, 500 pg/µl): 1 µlRecovery medium: 1 ml
StorageMSU440 ElectroCompetent 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 MSU440 ElectroCompetent 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 = 1×107
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.1274-12 1274-36
