Identification of toxin-binding protein involved in resistance to cry1 toxins, and related screening methods

2:

• The method of claim 1 wherein said method comprises running said sample on a gel, and assessing said alkaline phosphates enzymatic activity of an approximately 68 kDa protein band on said gel.

3:

• The method of claim 1 wherein said enzymatic activity is measured optically.

4:

• The method of claim 1 wherein said Cry1 protein is a Cry1Ac protein.

5:

• The method of claim 1 wherein said sample comprises brush border membrane vesicles (BBMV).

6:

• The method of claim 1 wherein said sample and said preparation comprise whole midguts from said lepidopteran and said insect.

7:

• The method of claim 1 wherein said lepidoptera is of the family Noctuidae.

8:

• The method of claim 1 wherein said lepidopteran is a Heliothis.

9:

• The method of claim 1 wherein said lepidopteran is a Heliothis virescens.

10:

• The method of claim 1 wherein said Cry1 protein binds an alkaline phosphatase insect receptor.

11:

• A kit for screening an insect for resistance to an insecticidal crystal protein wherein said kit comprises a collection means for obtaining a gut cell membrane sample from said insect, a holding means for holding said sample, and an assaying means for assaying said sample for an altered level of an alkaline phosphatase protein, wherein an altered level of alkaline phosphatase indicates a resistant insect and an unaltered level indicates a susceptible insect.

12:

• The kit of claim 11 where said assaying means is selected from the group consisting of a detectable antibody that binds an alkaline phosphatase, a detectable substrate that undergoes enzymatic degradation in the presence of alkaline phosphatase, and a nucleic acid probe.

13:

• The kit of claim 12 wherein said alkaline phosphatase protein is a receptor for a crystal protein insect toxin and said assaying means is an antibody and said antibody is a labeled antibody that specifically binds said alkaline phosphatase protein receptor.

14:

• The kit of claim 11 wherein said insect is a Heliothis virescens and said sample is assayed for a reduced amount of alkaline phosphatase activity.

15:

• A kit for performing the method of claim 1, wherein said kit comprises a collection means for obtaining a gut cell membrane sample from said insect, a holding means for holding said sample, and an assaying means for assaying said sample for an altered level of an alkaline phosphatase protein, wherein an altered level of alkaline phosphatase indicates a resistant insect and an unaltered level indicates a susceptible insect.

16:

• The kit of claim 15 where said assaying means is a detectable substrate that undergoes enzymatic degradation in the presence of alkaline phosphatase.

17:

• The kit of claim 15 wherein said insect is a Heliothis virescens and said sample is assayed for a reduced amount of alkaline phosphatase activity.