The LBA4404 strain is useful for transgenic operations of tomatoes, tobacco and other plants.
Intact Genomics LBA4404 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 LBA4404 strain is useful for transgenic operations of tomatoes, tobacco and other plants. LBA4404 contains a rifampicin resistance gene (rif) and also has streptomycin resistance. LBA4404 strain also contains a octoprine-type Ti plasmid pAL4404 without self-transport function, which contains the vir gene.
Competent cell type: Electrocompetent
Species: A. tumefaciens
Transformation efficiency: ≥ 1 x 107 cfu/µg pCAMBIA1391z DNA
Blue/white screening: No
Shipping condition: Dry ice
Reagents Needed for One Reaction
LBA4404 ElectroComp Agrobacterium: 25 µl
DNA (pCAMBIA1391z, 500 pg/µl): 1 µl
Recovery medium: 1 ml
LBA4404 ElectroComp Agrobacterium: -80 ºC
pCAMBIA1391z control DNA: -20 ºC
Recovery medium: 4 ºC
Transformation 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.
Follow these guidelines when using LBA4404 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 Efficiency
Transformation 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) 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 = 500
µg of DNA = 0.0005
Dilution = 100/1000 = 0.1
TE = 500/.0005/.1 = 1×1071285-12 1285-36