Weevil active proteins were identified previously with efficacy against the two African weevil species, Cylas puncticollis and C. brunneus, responsible of huge production loss of the sweetpotato in sub Saharan Africa. Transformation of sweetpotato with their respective genes (cry7Aa1 and cry3Ca1) yielded a total of 17 events with either cry gene. In order to identify events that are high expressers of the Cry protein, we tested transcriptional activity through real-time PCR based on SYBR Green I methodology and DAS-ELISA detection of the expressed protein using rabbit polyclonal antibodies. Five and nine transformed events bearing the cry7Aa1 and cry3Ca1 genes, respectively, were analyzed for transgene expression in the leaf, taking as reference the event with the lowest expression (LE). Two cry7Aa1 events showed statistically significant differences from the LE, with expression levels of 10.19 and 1.78 higher than the LE. Likewise, two cry3Ca1 events showed statistically significant differences from the LE, with expression levels of 54.65 and 2.78 times higher than the LE. These results were confirmed by DAS-ELISA analysis for protein expression in the leaves. The two cry7Aa1 events had protein expression levels of 0.03 and 0.009 ug/gr of fresh weight, and the two cry3Ca1 events had protein expression levels of 0.04 and 0.03 ug/gr of fresh weight. All other events had very low protein expression levels. Because the promoters used to guide gene expression were taken from the genes of the two major proteins in the tuberous root, it is expected that gene expression in the tuberous roots will be higher. This will allow us to localize the expression, a strategy used for insect resistance management.
Authors: Milagros Ormachea, Katterinne Prentice, Jose Carlos Tovar, Marc Ghislain, Milagros Ormachea, Katterinne Prentice, Jose Carlos Tovar, Marc Ghislain
Contributors: Jane Maurine Gati, Jane Maurine Gati
Publication Date: June2010
Keywords: African weevil, Cry genes, Sub Saharan Africa (SSA)
HOW TO CITE
Ormachea, M., Prentice, K., Tovar, J. and Ghislain, M. 2010. Expression of Sweet Potato-like Cry Genes to Control Weevils. In IN VITRO CELLULAR and DEVELOPMENTAL BIOLOGY-ANIMAL (Vol. 46, pp. S29-S29). 233 SPRING ST, NEW YORK, NY 10013 USA: SPRINGER.