AGL-1 (pSoup-P19) Agrobacterium Electrocompetent Cells

Product Description

AGL-1 (pSoup-P19) electrocompetent Agrobacterium cells are a highly efficient system for genetic transformation in various plant species. This strain is derived from the C58 RecA-type Agrobacterium strain and carries the rifampicin, carbenicillin, and streptomycin resistance genes for selection purposes. The Ti plasmid pTiBo542DT-DNA used in this system has a deleted T-DNA region, and instead contains a binary vector that restores the T-DNA binary system function.

The presence of the pSoup helper plasmid is required for the replication of pGreen, 62SK, and pGs2 series plasmids. The p19 protein is derived from tomato bush dwarf virus and further improves the stability of heterologous gene transcripts by inhibiting RNA silencing of foreign. These features help to enhance the Agrobacterium-mediated transformation of dicotyledonous plants like Arabidopsis thaliana, tobacco, and potato, as well as monocotyledonous plants such as corn.

Kit Components

  • Competent Cells
  • 1 x 12 mL Recovery Media
  • 1 x 10 µL Control Plasmid (pCAMBIA1391z Control, 500 pg/µL)
Product Specifications

Competent cell type: ElectroCompetent
Species: A. tumefaciens
Strain: AGL-1 (pSoup-P19)
Transformation efficiency: ≥1 x 108 cfu/µg pCAMBIA1391z DNA
Blue/white screening: No

Storage/Handling: This product may be shipped on dry ice. AGL-1 (pSoup-P19) 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.

Reagents Needed for One Reaction
  • AGL-1 (pSoup-P19) ElectroCompetent Agrobacterium: 25 µL
  • DNA (pCAMBIA1391z Control, 500 pg/µL): 1 µL
  • Recovery medium: 1 mL

Cuvette Offer Image-01

Table 1: Antibiotic disc sensitivity for GoldBio’s AGL-1 Agrobacterium strains (using standard BD antibiotic discs)


Antibiotic Selection

Amp

Carb

Chlor

Gent

Kan

Rif

Spect

Strep

Tet

100
µg/ml

100
µg/ml

30
µg/ml

100
µg/ml

30
µg/ml

50
µg/ml

5
µg/ml

50
µg/ml

50
µg/ml

5
µg/ml

AGL-1

R

R

R

n/a

R/S

S

R

S

R

S

AGL-1
(pSoup)

R

R

R

n/a

R/S

S

R

S

R

R

AGL-1
(pSoup-P19)

R

R

R

n/a

R/S

S

R

S

R

R

S = Sensitive
R = Resistant
R/S = intermediate zones using standard discs.
I = growth in inhibitory zone with standard disc. “Opaque”, not clear zone of inhibition.

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 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

pCAMBIA Plasmid Vector

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