Method capable of increasing competency of bacterial cell transformation

• US 6,709,854 B2
• Filed: 07/02/2001
• Issued: 03/23/2004
• Est. Priority Date: 02/02/1996
• Status: Expired due to Term

• Current Assignee: Invitrogen Corp. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Invitrogen Corp. (Thermo Fisher Scientific Incorporated)
• Inventors: Bebee, Robert L., Donahue, Robert A. Jr.
• Primary Examiner(s): Nashed, Nashaat T.
• Application Number:
  US09/895,202
• Publication Number:
  US 20010046698A1
• Time in Patent Office: 995 Days
• Field of Search: 435/252.8, 435/252.33, 435/440, 435/476, 435/488
• US Class Current: 435/252.33
• CPC Class Codes: C12N 1/20 Bacteria; Culture media therefor
• CPC Class Codes: C12N 15/10 Processes for the isolation, preparation or purification of DNA or RNA chemical preparation of DNA or RNA C07H21/00; preparation of non-structural polynucleotides from microorganisms or with enzymes C12P19/34
• CPC Class Codes: C12N 15/70 Vectors or expression systems specially adapted for E. coli

Abstract

The invention concerns bacterial strains capable of enhanced transformation efficiencies that are produced by the introduction of the F′ genetic material. The invention also concerns processes for producing transformable competent bacteria with enhanced transformation efficiencies.

Claims

1. One or more isolated E. coli cells wherein said cells are derived from DH5α

and comprise F′

genetic material derived from E. coli XL1-Blue or E. coli XL2-Blue, wherein said E. coli cells exhibit a higher transformation efficiency than said E. coli cells that do not contain F′

genetic material.

2. The E. coli cells of claim 1, further comprising exogenous DNA.

3. The E. coli cells of claim 2, wherein said exogenous DNA is plasmid DNA.

4. The E. coli cells of claim 2, wherein said exogenous DNA is lambda DNA.

5. The E. coli cells of claim 1, wherein said cells exhibit a transformation efficiency of 10⁸ or greater transformed bacteria per μ

g of purified plasmid DNA.

6. The E. coli cells of claim 5, wherein said cells exhibit a transformation efficiency of 10⁹ or greater transformed bacteria per μ

g of purified plasmid DNA.

7. One or more E. coli cells obtained by mating E. coli DH5α

cells with E. coli XL1-Blue cells or E. coli XL2-Blue cells.

8. The E. coli cells of claim 7, wherein said cells are obtained by mating E. coli DH5α

cells with E. coli XL1-Blue cells.

9. The E. coli cells of claim 7, wherein said cells are obtained by mating E. coli DH5α

cells with E. coli XL2-Blue cells.

10. A composition comprising:

(a) one or more E. coli cells, wherein said cells are derived from DH5α and

comprise F′

genetic material derived from E. coli XL1-Blue or E. coli XL2-Blue, wherein said E. coli cells exhibit a higher transformation efficiency than said E. coli cells that do not contain F′

genetic material; and

(b) a medium which supports the growth of said E. coli cells.

11. The composition of claim 10, further comprising one or more multivalent cations.

12. The composition of claim 11, wherein said one or more multivalent cations are magnesium cations.

13. A composition comprising:

(a) one or more E. coli cells wherein said cells are derived from DH5α and

comprise F′

genetic material derived from E. coli XL1-Blue or E. coli XL2-Blue, wherein said E. coli cells exhibit a higher transformation efficiency than said E. coli cells that do not contain F′

genetic material; and

(b) a solution capable of inducing competence in E. coli.

14. The composition of claim 13, wherein said solution capable of inducing competence in E. coli comprises one or more ingredients selected from the group consisting of:

potassium acetate, potassium chloride, manganese (II) chloride tetrahydrate, calcium chloride dihydrate, hexaminecobalt (III) chloride, glycerol and sucrose.

15. The composition of claim 14, wherein said solution capable of inducing competence in E. coli comprises 10 mM potassium acetate, 100 mM potassium chloride, 45 mM manganese (II) chloride tetrahydrate, 10 mM calcium chloride dihydrate, 3 mM hexaminecobalt (III) chloride, 10% glycerol and 5% sucrose.

16. The composition of claim 13, further comprising dimethylsulfoxide (DMSO).

17. The composition of claim 13, further comprising exogenous plasmid DNA.