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Submitted to: BARC Poster Day
Publication Type: Abstract Only Publication Acceptance Date: 5/18/1998 Publication Date: N/A Citation: N/A Interpretive Summary: Technical Abstract: Digestive proteinases of insects catalyze the release of free amino acids from dietary protein and thereby provide a supply of nutrients essential for normal growth and development. A possible approach to insect control is to express proteinase inhibitor (PI) genes in transgenic plants. Plant PIs have been shown in artificial feeding bioassays, as well as in transgenic plants, to inhibit gut proteinases and interfere with normal growth and development of insects. However, effective protection is likely to require multiple PIs directed against individual proteolytic activities in the insect gut. This problem is apparent in the growing number of instances in which insects exposed to a particular PI compensate by producing proteinases insensitive to that PI. Thus, the objective here is to identify multiple proteinase genes that may be involved in digestion. This will allow us to pursue an insect control strategy in which specific gut proteinases are targeted by PIs that are both specific and potent. To amplify cysteine proteinase genes, degenerate primer mixtures corresponding to evolutionarily conserved regions of amino acids within the enzymes were synthesized. These primers were used in PCR to amplify the corresponding region of the proteinase genes from a genomic DNA template. Templates included DNA from Hypera postica, as well as DNA from the dipteran insects Tetanops myopaeformis (sugarbeet root maggot) and Drosophila melanogaster (fruit fly) as positive control. Fragments of about 500bp have been amplified from each of these templates. Subcloning followed by selection, propagation, and sequencing of individual clones derived from each template can be expected to reveal one or more cysteine proteinase genes for each organism. Results will be presented. |
