COMBINATIONS INCLUDING CRY34AB/35AB AND CRY3Aa PROTEINS TO PREVENT DEVELOPMENT OF RESISTANCE IN CORN ROOTWORMS (DIABROTICA SPP.)

Inactive Publication Date: 2013-06-27
DOW AGROSCIENCES LLC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0017]The subject invention relates in part to Cry34Ab/35Ab in combination with Cry3Aa. The subject invention relates in part to the surprising discovery that Cry34Ab/Cry35Ab and Cry3Aa are useful for preventing development of resistance (to either insecticidal protein system alone) by a corn rootworm (Diabr

Problems solved by technology

Insects eat and damage plants and thereby undermine these human efforts.
Billions of dollars are spent each year to control insect pests and additional billions are lost to the damage they inflict.
Damage caused by insect pests is a major factor in the loss of the world's corn crops, despite the use of protective measures such as chemical pesticides.
Lodging eventually reduces corn yield and often results in death of the plant.
By fee

Method used

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  • COMBINATIONS INCLUDING CRY34AB/35AB AND CRY3Aa PROTEINS TO PREVENT DEVELOPMENT OF RESISTANCE IN CORN ROOTWORMS (DIABROTICA SPP.)
  • COMBINATIONS INCLUDING CRY34AB/35AB AND CRY3Aa PROTEINS TO PREVENT DEVELOPMENT OF RESISTANCE IN CORN ROOTWORMS (DIABROTICA SPP.)

Examples

Experimental program
Comparison scheme
Effect test

example 1

Expression and Purification

[0092]Construction of expression plasmids encoding Cry34Ab1, Cry35Ab1, and Cry3Aa full-length toxins. Standard cloning methods were used in the construction ofPseudomonas fluorescens (Pf) expression plasmids engineered to produce a full-length Cry34Ab1, Cry35Ab1, and Cry3Aa1 Cry proteins respectively. Restriction endonucleases from New England BioLabs (NEB; Ipswich, Mass.) were used for DNA digestion and T4 DNA Ligase from Invitrogen was used for DNA ligation. Plasmid preparations were performed using the Plasmid Midi Kit (Qiagen), following the instructions of the supplier. DNA fragments were purified using the Millipore Ultrafree®-DA cartridge (Billerica, Mass.) after agarose Tris-acetate gel electrophoresis. The basic cloning strategy entailed subcloning the coding sequences (CDS) of full-length Cry34Ab1, Cry35Ab1, and Cry3Aa1 Cry proteins into pMYC1803 at, for example, SpeI and XhoI (or XbaI, or HindIII) restriction sites, respectively, whereby they we...

example 2

Binding Assays

[0100]BBMV preparations. Brush border membrane vesicle (BBMV) preparations of insect midguts have been widely used for Cry toxin receptor binding assays. The BBMV preparations used in this invention were prepared from isolated midguts of third instars of the western corn rootworm (Diabrotica virgifera virgifera LeConte) using the method described by Wolferberger et al. (1987). Leucine aminopeptidase was used as a marker of membrane proteins in the preparation and Leucine aminopeptidase activities of crude homogenate and BBMV preparation were determined as previously described (Li et al. 2004a). Protein concentration of the BBMV preparation was measured using the Bradford method (1976).

[0101]125I Labeling. Purified full-length Cry34Ab1, chymotrypsinized Cry35Ab1, and trypsinized Cry3Aa were labeled using 125I for homologous and competition binding assays. To ensure the radio-labeling does not abolish the biological activity of the Cry toxins, cold iodination was conduct...

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Abstract

The subject invention relates in part to Cry34Ab/35Ab in combination with Cry3Aa. The subject invention relates in part to the surprising discovery that combinations of Cry34Ab/Cry35Ab and Cry3Aa are useful for preventing development of resistance (to either insecticidal protein system alone) by a corn rootworm (Diabrotica spp.) population. As one skilled in the art will recognize with the benefit of this disclosure, corn plants producing these insecticidal Cry proteins will be useful to mitigate concern that a corn rootworm population could develop that would be resistant to either of these insecticidal protein systems alone. Plants (and acreage planted with such plants) that produce these two insecticidal protein systems are included within the scope of the subject invention.

Description

BACKGROUND[0001]Humans grow corn for food and energy applications. Corn is an important crop. It is an important source of food, food products, and animal feed in many areas of the world. Insects eat and damage plants and thereby undermine these human efforts. Billions of dollars are spent each year to control insect pests and additional billions are lost to the damage they inflict.[0002]Damage caused by insect pests is a major factor in the loss of the world's corn crops, despite the use of protective measures such as chemical pesticides. In view of this, insect resistance has been genetically engineered into crops such as corn in order to control insect damage and to reduce the need for traditional chemical pesticides.[0003]Over 10 million acres of U.S. corn are infested with corn rootworm species complex each year. The corn rootworm species complex includes the northern corn rootworm (Diabrotica barberi), the southern corn rootworm (D. undecimpunctata howardi), and the western co...

Claims

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Application Information

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IPC IPC(8): C12N15/82B65D85/00A01G1/00A01N37/18
CPCC07K14/325C12N15/8286B32B1/02A01N37/18Y10T428/13A01G1/001A01H5/10A01N43/50B65D85/00A01H5/00C12N5/04Y02A40/146C12N15/8251A01N65/44A01G7/06
Inventor NARVA, KENNETH E.MEADE, THOMASFENCIL, KRISTIN J.LI, HUARONGHEY, TIMOTHY D.WOOSLEY, AARON T.OLSON, MONICA BRITT
Owner DOW AGROSCIENCES LLC
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