AGL-1 Agrobacterium Electrocompetent Cells
GoldBio’s AGL-1 Agrobacterium Electrocompetent Cells are optimized to produce the highest transformation efficiency, and are ideal for applications requiring high transformation efficiencies such as cDNA or gDNA library construction. The AGL-1 strain has a C58 chromosomal background that carries an insertion mutation in its recA recombination gene which stabilizes recombinant plasmids. It also carries rifampicin and carbenicillin resistance in its genome for selection. AGL-1 contains the Ti plasmid pTiBo542 from which the T-DNA region sequences have been deleted. Transformation with a binary vector containing the missing T-region results in a functional T-DNA binary system that allows for transfer of genetic material into a host plant’s genome. Therefore, this system is often used for Agrobacterium-mediated transformation of Arabidopsis thaliana as well as maize and other monocots.
Note: GoldBio agrobacterial strains have not
been comprehensively studied for antibiotic resistance. We recommend using
kanamycin for selection against agrobacterial strains, but it remains the researcher’s
responsibility to make sure that vectors are compatible with the agrobacterial
strains for alternative antibiotic selection.
Competent cell type: ElectroCompetent
Species: A. tumefaciens
Transformation efficiency: ≥1 x 107 cfu/µg pCAMBIA1391z DNA
Blue/white screening: No
Storage/Handling: This product may be shipped on dry ice. AGL-1 Agrobacterium Electrocompetent cells should be stored at -80°C, pCAMBIA1391z 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.
- ≥1 x 107 cfu/µg efficiency with electroporation.
Reagents Needed for One Reaction
- AGL-1 ElectroCompetent Agrobacterium: 25 µl
- DNA (pCAMBIA1391z, 100 pg/µl): 1 µl
- Recovery medium: 1 ml
Transformation efficiency is tested by using the pCAMBIA1391z control DNA supplied with the kit and using the protocol given below. Transformation efficiency should be ≥6 x 107 CFU/µg pCAMBIA1391z DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.
- 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 capable of generating field strengths of 16 kV/cm is required.
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
Procedure for transformation through electroporation utilizing ALG-1 Agrobacterium Electrocompetent Cells.
Guide to GoldBio's competent cell collection containing transformation efficiencies, characteristics and applications for each of our competent cell lines.
Chart containing genotypes and transformation efficiencies for GoldBio's competent cell collection.
This guide provides nomenclature information relating to genotypes and genetic markers of E. coli and competent cells.