Intact Genomics (ig^®) Methionine-Auxotrophic EHA105^Met Chemically Competent Agrobacterium cells include modifications so that they will not grow unless free methionine is added to Minimal medium.  This prevents the bacteria from overgrowing plant tissues when used for plant transformation.  Methionine- Auxotrophic EHA105^Met Chemically Competent Agrobacterium cells are optimized for the highest transformation efficiencies. The EHA105^Met strain is useful for transgenic operations of rice, tobacco and other plants. EHA105 contains a rifampicin resistance gene (rif). EHA105^Met strain also contains an amber basic Ti plasmid pEHA105 (pTiBo542DT-DNA) without self-transport function, which contains the vir gene.

Custom Aliquots Available

Benefits

• Methionine Auxotrophic
• Enables development of more efficient transformation systems
• Reduced bacterial overgrowth during co-cultivation
• Decreased need for antibiotics

Specifications

Competent cell type:  Chemically CompetentSpecies:  A. tumefaciensFormat: TubesTransformation efficiency:  ≥ 1 x 10⁵ cfu/µg pUC19 DNAShipping condition:  Dry ice

Reagents Included

• ig^® Methionine-Auxotrophic EHA105^Met Chemically Competent Agrobacterium Cells
• DNA (pCAMBIA1391z, 500 pg/µl)
• Recovery medium

Storage

• ig^® Methionine Auxotrophic EHA105^Met Chemically Competent Agrobacterium Cells: -80 ºC
• pCAMBIA1391z control DNA: -20 ºC
• Recovery medium: 4 ºC

Quality Control

Transformation efficiency is tested by using the pCAMBIA1391z control DNA supplied with the kit and using the protocol given below. Transformation efficiency should be ≥1 x 10⁵ CFU/µg pCAMBIA1391z DNA. Untransformed cells are tested for appropriate antibiotic sensitivity. 

General Guidelines

Follow these guidelines when using ig^® Methionine-Auxotrophic EHA105^Met Chemically Competent 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.

Example 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/Dilution

Transform 1 µl of (10 pg/µl) pUC19 control plasmid into 50 µl of cells, add 950 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 100 colonies, the TE is calculated as follows:

Colonies = 5µg of DNA = 0.0005Dilution = 100/1000=0.1TE = 5/.0005/.1 = 1.0×10⁵1078-05 1078-15