Description
GoldBio’s GV3101 Agrobacterium
Electrocompetent Cells allow you to obtain high transformation
efficiency in applications such as gDNA or cDNA library construction.
Our GV3101 strain harbors the C58 chromosomal backbone containing
rifampicin resistance and the Ti plasmid pmp90 (pTiC58DT-DNA) harboring
the gentamicin resistance. A functional T-DNA binary system can be built
using our GV3101 strains as the T-DNA region has been deleted from the
Ti plasmid and instead has a binary vector containing the missing
T-region. The binary system makes possible to transfer genetic material
into a host plant’s genome. Our system is often used for
Agrobacterium-mediated transformation in mono and dicotyledonous species such as Arabidopsis thaliana, tobacco, potato, soybeans and corn.
Table 1: Antibiotic disc sensitivity for GoldBio’s Agrobacterium strains (using standard BD antibiotic discs)
|
Antibiotic Selection |
||||||||||
|
Amp |
Carb |
Chlor |
Gent |
Kan |
Rif |
Spect |
Strep |
Tet |
||
|
100 |
100 |
30 |
100 |
30 |
50 |
25 |
50 |
50 |
50 |
|
|
GV3101 |
I |
R |
R |
PR |
R |
S |
R |
S |
R |
S |
|
EHA 105 |
R |
R/S |
R |
n/a |
R/S |
S |
R |
S |
R |
S |
|
LBA 4404 |
S |
S |
S |
n/a |
S |
S |
R |
S |
R |
S |
|
AGL-1 |
R |
R |
R |
<p n/a |
R/S |
S |
R |
S |
R |
S |
|
C58C1 |
R |
R |
R |
n/a |
R/S |
S |
R |
S |
R |
S |
|
S = Sensitive I = |
||||||||||
Product Specifications
Competent cell type: ElectroCompetent
Species: A. tumefaciens
Strain: GV3101
Transformation efficiency: ≥2 x 108 cfu/µg pCAMBIA1391z DNA
Blue/white screening: No
Storage/Handling: This product may be shipped on dry ice. GV3101 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.
Genomic Features
- ≥2 x 108 cfu/µg efficiency with electroporation.
Reagents Needed for One Reaction
- GV3101 ElectroCompetent Agrobacterium: 25 µl
- DNA (pCAMBIA1391z, 500 pg/µl): 1 µl
- Recovery medium: 1 ml
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 ≥2 x 108 CFU/µg pCAMBIA1391z DNA. Untransformed cells are tested for appropriate antibiotic sensitivity.
General Guidelines
- 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
