Intact Genomics (ig^®) Ar.A4 Chemically Competent Agrobacterium cells are made from a specific strain of Rhizobium rhizogenes (formerly Agrobacterium rhizogenes), Agrobacterium rhizogenes(kan R) Ar A4 Ri(agropine type). Agrobacterium rhizogenes is a soil-borne gram-negative bacterium that can infect most dicotyledons, a few monocotyledons and some gymnosperms.  Ar.A4 Chemically Competent Agrobacterium are optimized for the highest transformation efficiencies and are useful for transgenic operations of corn, tobacco, carrot and other plants.  Ar.A4 Agrobacterium rhizogenes strain contains agrobacterium-type Ri plasmid and displays kanamycin resistance.

SpecificationsCompetent cell type: Chemically CompetentSpecies: R. rhizogenesStrain: Ar.A4Format: TubesTransformation efficiency:  ≥ 1 x 10⁵  cfu/µg pIG7spe DNABlue/white screening: NoShipping condition: Dry ice

Reagents Included

• Ar.A4 Chemically Competent Agrobacterium
• DNA (pIG7spe, 500 pg/µl)
• Recovery medium

Note: Liquid nitrogen is required.

StorageAr.A4 Chemically Competent Agrobacterium: -80 ºCpIG7spe control DNA: -20 ºCRecovery medium: 4 ºC

Quality ControlTransformation efficiency is tested by using the pIG7spe control DNA supplied with the kit and using the protocol in this manual. Transformation efficiency should be ≥1 x 10⁵ CFU/µg pIG7spe DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.

General Guidelines

Follow these guidelines when using Ar.A4 Chemically Competent Agrobacterium cells:

• 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 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/Plated

Transform 1 µl of (500 pg/µl) pIG7spe control plasmid into 50 µl of cells, add 950 µl of Recovery Medium. Recover for 3 hours and plate 100 µl. Count the colonies on the plate in two days. If you count 5 colonies, the TE is calculated as follows:

Colonies = 5µg of DNA = 0.0005Dilution = 100/1000 = 0.1TE = 5/.0005/.1 = 1×10⁵

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 spectinomycin-selection.1072-06 1072-18