FORMATION OF AND METHODS FOR THE PRODUCTION OF LARGE BACILLUS THURINGIENSIS CRYSTALS WITH INCREASED PESTICIDAL ACTIVITY

CIPO - PATENT - 2167178 http://patents.ic.gc.ca/cipo/cpd/en/patent/2167178/summary.html

NOTE ONE OF MANY (United States Patent 6,280,721): wherein said heterologous crystal endotoxin is produced from a homologously recombined gene in said integrant's chromosome, and wherein said integrant has greater pesticidal activity than a corresponding parental strain by producing a larger quantity of a crystal delta-endotoxin as compared to said corresponding parental strain. LOOKS LIKE A BIT OF CLONING HERE (?)

Patent Application: (11) CA 2167178

(54) FORMATION OF AND METHODS FOR THE PRODUCTION OF LARGE BACILLUS THURINGIENSIS CRYSTALS WITH INCREASED PESTICIDAL ACTIVITY

(54) FORMATION ET METHODES DE PRODUCTION DE GRANDS CRISTAUX DE BACILLUS THURINGIENSIS, POSSEDANT UNE PLUS GRANDE ACTIVITE PESTICIDE

ABSTRACT

The invention relates to a method for producing an integrant(s) of Bacillus thuringiensis which produces a larger quantity of a crystal delta-endotoxin with greater pesticidal activity as compared to the crystal delta-endotoxin produced by the corresponding parental strain. The crystal delta-endotoxin produced by the integrant Bacillus thuringiensis will have an activity directed towards the same pest(s) as its parent Bacillus thuringiensis crystal delta-endotoxin. The invention further relates to such integrants, compositions comprising such integrants, as well as methods for controlling a pest(s) using these compositions.

CLAIMS Show all claims

*** Note: Data on abstracts and claims is shown in the official language in which it was submitted.

(72) Inventors: (Country) ADAMS, LEE FREMONT (United States)

THOMAS, MICHAEL DAVID (United States)

SLOMA, ALAN P. (United States)

WIDNER, WILLIAM R. (United States)

(73) Owners: (Country) ABBOTT LABORATORIES (United States)

(71) Applicants: (Country)

(74) Agent: OGILVY RENAULT LLP/S.E.N.C.R.L.,S.R.L.

(45) Issued:

(86) PCT Filing Date: 1994-07-14

(87) PCT Publication Date: 1995-01-26

Examination requested: 2001-06-12

(51) International Class (IPC): C12N 15/75 (2006.01)

A01N 37/18 (2006.01)

A01N 43/36 (2006.01)

A01N 43/50 (2006.01)

A01N 63/00 (2006.01)

C07K 14/325 (2006.01)

C12N 1/21 (2006.01)

C12N 15/32 (2006.01)

Patent Cooperation Treaty (PCT): Yes

(86) PCT Filing Number: PCT/US1994/007955

(87) International publication number: WO1995/002695

(85) National Entry: 1996-01-12

(30) Application priority data:

Application No. Country Date

08/092,338 United States 1993-07-15

Availability of licence: N/A

Language of filing: English

United States Patent 6,303,382

Adams , et al. October 16, 2001

Formation of and methods for the production of large bacillus thuringiensis crystals with increased pesticidal activity

Abstract

Inventors: Adams; Lee Fremont (Davis, CA), Thomas; Michael David (Davis, CA), Sloma; Alan P. (Davis, CA), Widner; William R. (Davis, CA)

Appl. No.: 08/771,190

Filed: December 20, 1996

This application is a divisional of U.S. patent application Ser. No. 08/274,608, filed Jul. 13, 1994, which is a continuation-in-part of U.S. patent application Ser. No. 08/092,338, filed Jul. 15, 1993, now abandoned.

United States Patent 6,280,721

Adams , et al. August 28, 2001

Production of Bacillus thuringiensis integrants

Abstract

The invention relates to a method for producing an integrant(s) of Bacillus thuringiensis. The invention further relates to such integrants, compositions comprising such integrants, as well as methods for controlling a pest(s) using these compositions.

Inventors: Adams; Lee Fremont (Davis, CA), Thomas; Michael David (Davis, CA), Sloma; Alan P. (Davis, CA), Widner; William R. (Davis, CA), J.o slashed.rgensen; Steen Troels (Aller.o slashed.d, DK), J.o slashed.rgensen; Per Lin.ang. (Copenhagen, DK), Diderichsen; B.o slashed.rge Krag (Birker.o slashed.d, DK)

Assignee: Valent BioSciences, Inc. (Libertyville, IL)

Appl. No.: 08/377,891

Filed: January 25, 1995

This application is a continuation-in-part of co-pending application serial no. 08/274,608, filed Jul. 13, 1994, which is a continuation-in-part of application Ser. No. 08/092,338, filed Jul. 15, 1993, now abandoned. This application is also a continuation-in-part of application Ser. No. 07/853,701, filed May 26, 1992.

What is claimed is:

1. An integrant of Bacillus thuringiensis or spore thereof which produces at least one heterologous crystal delta-endotoxin, wherein said heterologous crystal endotoxin is produced from a homologously recombined gene in said integrant's chromosome, and wherein said integrant has greater pesticidal activity than a corresponding parental strain by producing a larger quantity of a crystal delta-endotoxin as compared to said corresponding parental strain.

2. The integrant of claim 1 in which the integrant is an integrant of Bacillus thuringiensis subsp. kurstaki.

3. The integrant of claim 1 in which the integrant is an integrant of a cry.sup.- strain.

4. The integrant of claim 1 in which the heterologous delta-endotoxin is a CryIC protein.

5. The integrant of claim 1 in which the integrant has all of the identifying characteristics of strain EMCC0122, deposited with the NRRL, having an accession number of NRRL B-21386.

6. The integrant according to claim 1 in which the delta-endotoxin produced is active against an insect pest.

7. The integrant according to claim 1 in which the delta-endotoxin produced is active against an insect pest of the order Lepidoptera.

8. A pesticidal composition comprising (a) the integrant of claim 1 and (b) a pesticidally acceptable carrier.

9. A DNA construct comprising (a) an origin of replication but lacking a functional gene or portion thereof encoding a factor required for plasmid replication from the origin of replication; (b) a DNA sequence encoding a Bacillus thuringiensis delta-endotoxin; (c) a DNA sequence that is homologous with a region of the genome of a Bacillus thuringiensis; and (d) a selectable marker.

10. A DNA vector comprising the DNA construct of claim 9.

_________________

United States Patent 6,280,720

Adams , et al. August 28, 2001

Formation of and methods for the production of large Bacillus thuringiensis crystals with increased pesticidal activity

Abstract

The invention relates to a method for producing an integrant(s) of Bacillus thuringiensis which produces a larger quantity of a crystal delta-endotoxin greater pesticidal activity as compared to the crystal delta-endotoxin produced by the corresponding parental strain. The crystal delta-endotoxin produced by the integrant Bacillus thuringiensis will have an activity directed towards the same pest(s) as its parent Bacillus thuringiensis crystal delta-endotoxin. The invention further relates to such integrants, compositions comprising such integrants, as well as methods for controlling a pest(s) using these compositions.

Inventors: Adams; Lee Fremont (Davis, CA), Thomas; Michael David (Davis, CA), Sloma; Alan P. (Davis, CA), Widner; William R. (Davis, CA)

Assignee: Valant BioSciences, Inc. (

Libertyville, Inc. (

Appl. No.: 08/274,608

Filed: July 13, 1994

This application is a continuation-in-part application of application Ser. No. 08/092,338 filed Jul. 15, 1993, abandoned, the contents of which are incorporated herein by reference.

What is claimed is:

1. An integrant of Bacillus thuringiensis or spore thereof which produces at least one homologous or heterologous crystal delta-endotoxin, wherein said homologous or heterologous crystal delta-endotoxin is produced form a homologously recombined gene in said integrant's chromosome, and wherein said integrant has greater pesticidal activity than a corresponding parental strain by producing a larger quantity of a crystal delta-endotoxin as compared to said corresponding parental strain.

2. The integrant according to claim 1, wherein the delta-endotoxin produced is active against an insect pest.

3. The integrant according to claim 1, wherein the delta-endotoxin produced is active against an insect pest selected from the group consisisting of an insect pest of the order Coleoptera and an insect pest of the order Lepidoptera.

4. A pesticidal composition comprising (a) the integrant of claim 1 and (b) a pesticidally acceptable carrier.

5. A method for controlling a pest comprising exposing the pest to a pest-controlling effective amount of the pesticidal composition of claim 4.

6. A DNA construct lacking a Bacillus thuringiensis origin of replication comprising (i) a DNA sequence encoding a delta-endotoxin wherein said DNA sequence is obtained from a Bacillus thuringiensis strain producing larger amounts of said delta-endotoxin than a parental strain producing said delta-endotoxin; (ii) a DNA sequence that is homologous with a region of the genome of a Bacillus cell; and (iii) a selectable marker.

7. A recombinant DNA vector comprising the DNA construct of claim 6.

______________

United States Patent 6,270,760

Adams , et al. August 7, 2001

Production of Bacillus thuringiensis integrants

Abstract

Assignee: Valent BioSciences, Inc. (Libertyville, IL)

Appl. No.: 08/872,571

Filed: June 10, 1997

This application is a Continuation of application Ser. No. 08/377,892, filed Jan. 25, 1995, now abandoned which is a continuation-in-part of co-pending application Ser. No. 08/274,608, filed Jul. 13, 1994, which is a continuation-in-part of application Ser. No. 08/092,338, filed Jul. 15, 1993, now abandoned. This application is also a continuation-in-part of application Ser. No. 07/853,701, filed May 26, 1992 now abandoned.

Claims

What is claimed is:

1. A method for controlling a pest, the method comprising the step of exposing the pest to a pest-controlling effective amount of a pesticidal composition comprising (a) an integrant of Bacillus thuringiensis or spore thereof which produces at least one heterologous crystal delta-endotoxin in said integrant, wherein said integrant is produced by a method comprising the steps of: (i) introducing into a cell of a host Bacillus thuringiensis strain a first DNA vector comprising a first origin of replication and at least one functional gene encoding at least one factor required for plasmid replication from said first origin of replication, a second DNA vector comprising a second origin of replication but lacking a functional gene encoding a factor required for plasmid replication from the second origin of replication and a DNA sequence encoding a Bacillus thuringiensis delta-endotoxin, a DNA sequence homologous with a region of the genome of said host strain, and a selectable marker and (ii) culturing the cell of step (i) under selective conditions leading to the loss of the first DNA vector and integration of the second DNA vector into the genome of the host cell by homologous recombination, and (b) a pesticidally acceptable carrier.

2. The method of claim 1 in which the integrant is an integrant of Bacillus thuringiensis subsp. kurstaki.

3. The method of claim 1 in which the integrant is an integrant of a cry- strain.

4. The method of claim 1 in which the heterologous delta-endotoxin is a CryIC protein.

5. The method of claim 1 in which the integrant has all of the identifying characteristics of strain EMCC0122, deposited with the NRRL, having an accession number of NRRL B-21386.

6. The method of claim 1 in which the integrant has greater pesticidal activity than a corresponding parental strain by producing a larger quantity of a crystal delta-endotoxin as compared to said corresponding parental strain.

7. The method according to claim 1 in which the delta-endotoxin produced is active against an insect pest.

8. The method according to claim 1 in which the delta-endotoxin produced is active against an insect pest of the order Lepidoptera.

_________________________

United States Patent 5,955,367

Adams , et al. September 21, 1999

Production of bacillus thuringiensis integrants

Abstract

Inventors: Adams; Lee Fremont (Davis, CA), Thomas; Michael David (Davis, CA), Sloma; Alan P. (Davis, CA), Widner; William R. (Davis, CA), J.o slashed.rgensen; Steen Troels (Aller.o slashed.d, DK), J.o slashed.rgensen; Per Lin.ang. (Copenhagen K, DK), Diderichsen; B.o slashed.rge Krag (Birker.o slashed.d, DK)

Assignee: Abbott Laboratories (Abbott Park, IL)

Appl. No.: 09/060,288

Filed: April 14, 1998

This is a continuation of U.S. patent application Ser. No. 08/378,236, filed Jan. 25, 1995 now abandoned, which is a continuation-in-part of co-pending application Ser. No. 08/274,608, filed Jul. 13, 1994, which is a continuation-in-part of application Ser. No. 08/092,338, filed Jul. 15, 1993, now abandoned. This application is also a continuation-in-part of application Ser. No. 07/853,701, filed May 26, 1992, now abandoned, which claims benifit of PCT/DK90/00332, filed Dec. 18, 1990.

Claims

What is claimed is:

1. A method for obtaining an integrant of Bacillus thuringienis comprising:

(a) introducing into a cell of a host Bacillus thuringiensis strain (i) a first DNA vector comprising a first origin of replication and at least one function gene encoding at least one temperature sensitive replication protein required for plasmid replication from said first origin of replication, and (ii) a second DNA vector comprising a second origin of replication and a selectable marker but lacking a functional gene or portion thereof encoding a factor required for plasmid replication from the second origin of replication, as well as a heterologous DNA sequence encoding a Bacillus thuringinesis delta-endotoxin, and a DNA sequence that is homologous with a region of the genome of said parental strain, wherein said second vector cannot replicate in the absence of said first vector and

(b) culturing the cell of step (a) under selective conditions of temperature and antibiotic pressure leading to integration of said second DNA vector into the genome of said parental cell by homologous recombination and loss of the first DNA vector.

2. The method according to claim 1 in which the host Bacillus thuringiensis strain is a cry.sup.- strain.

3. The method according to claim 1 in which the host Bacillus thuringiensis strain is selected from the group consisting of Bacillus thuringiensis subsp. kurstaki, Bacillus thuringiensis subsp. aizawai, Bacillus thuringiensis subsp. galleriae, Bacillus thuringiensis subsp. entomocidus, Bacillus thuringiensis subsp. tenebrionis, Bacillus thuringiensis subsp. thuringiensis, Bacillus thuringiensis subsp. alesti, Bacillus thuringiensis subsp. canadiensis, Bacillus thuringiensis subsp. darmstadiensis, Bacillus thuringiensis subsp. dendrolimus, Bacillus thuringiensis subsp. finitimus, Bacillus thuringiensis subsp. kenyae, Bacillus thuringiensis subsp. morrisoni, Bacillus thuringiensis subsp. subtoxicus, Bacillus thuringiensis subsp. toumanoffi, Bacillus thuringiensis subsp. toumanoffi, Bacillus thuringiensis subsp. pondicheriensis, Bacillus thuringiensis subsp. shandogiensis, Bacillus thuringiensis subsp. sotto, Bacillus thuringiensis subsp. nigeriae, Bacillus thuringiensis subsp. yunnanensis, Bacillus thuringiensis subsp. dakota, Bacillus thuringiensis subsp. indiana, Bacillus thuringiensis subsp. tohokuensis, Bacillus thuringiensis subsp. kumamotoensis, Bacillus thuringiensis subsp. tochigiensis, Bacillus thuringiensis subsp. t hompsoni, Bacillus thuringiensis subsp. wuhanensis, Bacillus thuringiensis subsp. kyushuensis, Bacillus thuringiensis subsp. ostriniae, Bacillus thuringiensis subsp. tolworthi, Bacillus thuringiensis subsp. pakistani, Bacillus thuringiensis subsp japonensis, Bacillus thuringiensis subsp. colmeri, Bacillus thuringiensis subsp. pondicheriensis, Bacillus thuringiensis subsp. shandongiensis, Bacillus thuringiensis subsp. neoleonensis, Bacillus thuringiensis subsp. coreanensis, Bacillus thuringiensis subsp. silo, Bacillus thuringiensis subsp. mexcanensis, and Bacillus thuringiensis subsp. israelensis.

4. The method according to claim 1 in which the host Bacillus thuringiensis strain is Bacillus thuringiensis subsp. kurstaki.

5. The method according to claim 1 in which the DNA sequence encoding a delta-endotoxin is a cryIC gene.

6. The method according to claim 1 in which the DNA sequence encoding the delta-endotoxin is obtained from a Bacillus thuringiensis strain selected from the group consisting of Bacillus thuringiensis subsp. kurstaki, Bacillus thuringiensis subsp. aizawai, Bacillus thuringiensis subsp. galleriae, Bacillus thuringiensis subsp. entomocidus, Bacillus thuringiensis subsp. tenebrionis, Bacillus thuringiensis subsp. thuringiensis, Bacillus thuringiensis subsp. alesti, Bacillus thuringiensis subsp. canadiensis, Bacillus thuringiensis subsp. darmnstadiensis, Bacillus thuringiensis subsp. dendrolimus, Bacillus thuringiensis subsp. finitimus, Bacillus thuringiensis subsp. kenyae, Bacillus thuringiensis subsp. morrisoni, Bacillus thuringiensis subsp. subtoxicus, Bacillus thuringiensis subsp. toumanoffi, Bacillus thuringiensis subsp. toumanoffi, Bacillus thuringiensis subsp. pondicheriensis, Bacillus thuringiensis subsp. shandogiensis, Bacillus thuringiensis subsp. sotto, Bacillus thuringiensis subsp. nigeriae, Bacillus thuringiensis subsp. yunnanensis, Bacillus thuringiensis subsp. dakota, Bacillus thuringiensis subsp. indiana, Bacillus thuringiensis subsp. tohokuensis, Bacillus thuringiensis subsp. kumamotoensis, Bacillus thuringiensis subsp. tochigiensis, Bacillus thuringiensis subsp. thompsoni, Bacillus thuringiensis subsp. wuhanensis, Bacillus thuringiensis subsp. kyushuensis, Bacillus thuringiensis subsp. ostriniae, Bacillus thuringiensis subsp. tolworthi, Bacillus thuringiensis subsp. pakistani, Bacillus thuringiensis subsp. japonensis, Bacillus thuringiensis subsp. colmeri, Bacillus thuringiensis subsp. pondicheriensis, Bacillus thuringiensis subsp. shandongiensis, Bacillus thuringiensis subsp. neoleonensis, Bacillus thuringiensis subsp. coreanensis, Bacillus thuringiensis subsp. silo, Bacillus thuringiensis subsp. mexcanensis, and Bacillus thuringiensis subsp. israelensis.

7. The method according to claim 1 in which the DNA sequence encoding the delta-endotoxin is obtained from Bacillus thuringiensis subsp. aizawai.

8. The method according to claim 1 in which the selectable marker from the second DNA vector is a DNA sequence encoding antibiotic resistance.

9. The method according to claim 1 in which the first DNA vector further comprises a selectable marker and in which said selectable marker differs from the selectable marker in the second DNA vector.

10. The method according the claim 1, in which the first DNA vector comprises a first origin of replication from a single-strand DNA plasmid and a functional rep gene, and in which the second DNA vector comprises a second origin of replication from a single-strand DNA plasmid but lacking a functional rep gene, a DNA sequence encoding a Bacillus thuringiensis delta-endotoxin, and a DNA sequence that is homologous with a region of the genome of said host strain.

11. The method according to claim 1 in which the cell in step (b) is incubated at about 37.degree. C.

12. The method according to claim 1 in which said method further comprises amplifying the integrated DNA sequence by culturing the integrant of step (b) in the presence of increasing amounts of a selecting-agent.

13. The method according to claim 1 in which the integrant has all of the identifying characteristics of strain EMCC0122, deposited with the NRRL, having an accession number of NRRL B-21386.

PATENTS SHOWING "DELTA-ENDOTOXIN" (Agraquest and/or Agraquest Scientists

PAT. NO. Title

6,844,182

Publication number: US2002004237

Also published as:

US6844182 (B2)

international: A01N47/28; A01N63/00; A01N63/02; C07C275/16; C12P13/02; A01N47/28; A01N63/00; A01N63/02; C07C275/00; C12P13/00; (IPC1-7): A01N63/00; C12N1/20

- European: A01N47/28; A01N63/00; A01N63/02; C07C275/16; C12P13/02; C12R1/07

Application number: US20010858207 20010515

Priority number(s): US20010858207 20010515; US19980013375 19980126; US19950454967 19950530; US19930146852 19931103; US19930095240 19930720; US19920990202 19921214; US19920971786 19921105

Mutants which produce a potentiator of bacillus pesticidal activity.

This invention is directed to Bacillus mutants which produce a factor

which potentiates the pesticidal activity of a Bacillus related pesticide, a

chemical pesticide and/or a virus with pesticidal properties.

As defined herein, "a Bacillus related pesticide" is a Bacillus (e.g., Bacillus

thuringiensis or Bacillus subtilis) strain, spore, or substance, e.g., protein

or fragment thereof having activity against or which kill pests or a

microorganism capable of expressing a Bacillus gene encoding a Bacillus

protein or fragment thereof having activity against or which kill pests

(e.g., Bacillus thuringiensis delta-endotoxin) and an acceptable carrier

(see Section 5.2., infra, for examples of such carriers).

Inventors: Outtrup; Helle (Denmark, DK), Lidster; William D. (Sacramento,

CA), MacIntosh; Susan C. (Woodland, CA), Starnes; Robert L. (Sacramento,

CA), Manker; Denise Carol (Davis, CA)

Assignee: Valent BioSciences Corporation (Libertyville, IL)

Appl. No.: 09/858,207

Filed: May 15, 2001

6,268,181

Methods for producing a potentiator of Bacillus pesticidal activity.

This invention is directed to a method for obtaining a factor which

potentiates the pesticidal activity of a Bacillus related pesticide, a

chemical pesticide and/or a virus with pesticidal properties.

B. thuringiensis subsp. kurstaki strain EMCC0086 (NB75 deposited

with the NRRL as B-21147 ..

The isolated Ia is found to potentiate the activity of Bacillus

thuringiensis subsp. kurstaki and Bacillus thuringiensis subsp.

aizawai crystal delta-endotoxin pesticidal protein

Inventors: Manker; Denise Carol (Davis, CA), Lidster; William D.

(Sacramento, CA), MacIntosh; Susan C. (Woodland, CA), Starnes; Robert L.

(Sacramento, CA)

Assignee: Valent BioSciences, Corporation (Libertyville, IL)

Appl. No.: 09/015,647

Filed: January 28, 1998

6,210,952

Full-Text Bacillus thuringiensis mutants which

produce higher yields of crystal delta-endotoxin than their corresponding

parental strains

United States Patent 6,210,952

Liu , et al. April 3, 2001

Bacillus thuringiensis mutants which produce higher yields of crystal

delta-endotoxin than their corresponding parental strains

Abstract

The invention relates to a mutant of Bacillus thuringiensis which produces

a larger amount of crystal delta-endotoxin with a greater pesticidal

activity as compared to the corresponding parental strain. The mutant

may also have a larger crystal size as compared to the corresponding

parental strain. The crystal delta-endotoxin produced by the mutant

Bacillus thuringiensis will have an activity directed towards the same

pest(s) as its parental Bacillus thuringiensis crystal delta-endotoxin. The

invention further relates to a method for producing such a mutant,

compositions comprising such a mutant as well as methods for

controlling a pest(s) using these compositions.

Inventors: Liu; Chi-Li (Davis, CA); Marrone; Pamela Gail (Davis,

CA); Payne; Jewel M. (Davis, CA); Gurtler; Hanne (Holte, DK); Petersen;

Annette Schousboe (Birker.o slashed.d, DK)

Assignee: Valent BioSciences Corp. (Libertyville, IL)

Appl. No.: 902412

Filed: July 29, 1997

5,919,447

Full-Text Strain of bacillus [Bacillus

thuringiensis] for controlling plant disease

Inventors: Marrone; Pamela Gail (Davis, CA); Heins; Sherry D.

(Davis, CA); Manker; Denise C. (Davis, CA); Jimenez; Desmond R.

(Woodland, CA); Chilcott; Chris N. (Auckland, NZ); Wigley; Peter

(Auckland, NZ); Broadwell; Andrew (Auckland, NZ)

Assignee: Agraquest, Inc. (Davis, CA)

5,874,289

Full-Text Bacillus thuringiensis mutants which

produce high yields of crystal delta-endotoxin

United States Patent 5,874,289

Liu , et al. February 23, 1999

Bacillus thuringiensis mutants which produce high yields of crystal delta-

endotoxin