August 2011
Source: Crop Biotech Update
Chickpea (Cicer arietinum L.) is the second important legume planted across the globe next to soybeans. It is a major source of dietary protein both for human consumption and feed for livestock. Despite the huge demand for the crop, the global yield of chickpea is stagnating for the past twenty years due to various biotic and abiotic stresses that affect its growth.
One of the ways to improve the production of the crop is through the use of modern biotechnology techniques. Thus, Meenakshi Mehrotr of the National Botanical Research Institute in India, together with other researchers obtained stable transgenic chickpea using Agrobacterium-mediated transformation of cotyledonary nodes (CN) with dicot-preferred modified truncated synthetic cry1Ab and cry1Ac genes.
Based on the insect bioassay performed with the transgenic plants, there was relatively higher toxicity for plants with Cry1AC protein compared to Cry1Ab to pod borer insect (Helicoverpa armigera). Transgenic plants with medium expression of stacked Bt genes exhibited strong resistance and protection against pod borer larvae than plants with high expression of a single Bt gene. Results showed the importance of pyramiding genes and co-expression of Bt genes for effective protection against lepidopteran pests of chickpea.
Pyramiding of modified cry1Ab and cry1Ac genes of Bacillus thuringiensis in transgenic chickpea (Cicer arietinum L.) for improved resistance to pod borer insect Helicoverpa armigera
Meenakshi Mehrotra, Aditya K. Singh, Indraneel Sanyal, Illimar Altosaar and D. V. Amla
Abstract
The modified cry1Ab and cry1Ac insecticidal genes of Bacillus thuringiensis (Bt) under the control of two different constitutive promoters have been introduced into chickpea (Cicer arietinum L.) by Agrobacterium-mediated transformation of pre-conditioned cotyledonary nodes. 118 stable transformed T0 plants as independent transformation events were obtained expressing individual cry1Ab, cry1Ac or both pyramided genes for their co-expression driven by either cauliflower mosaic virus 35S promoter with duplicated enhancer (CaMV35S) or synthetic constitutive promoter (Pcec) and their combinations. Integration and inheritance of transgenes in T0 and T1 population of transgenic chickpea plants were determined by PCR, RT-PCR and Southern hybridization. Results of Southern hybridization showed single copy integration of cry1Ab or cry1Ac genes in most of the transgenic plants developed with either single or pyramided genes and reflected Mendelian inheritance of transgenes in T1 progeny. Real time PCR of pyramided transgenic plants clearly showed differential expression of transcripts for both the genes driven by CaMV35S and Pcec promoters. Quantitative assessment of Bt Cry toxins by ELISA of T0 transgenic chickpea plants showed expression of toxin ranging from 5 to 40 ng mg−1 of total soluble protein (TSP) in leaves of transgenic plants. Insect bioassay performed with transgenic plants showed relatively higher toxicity for plants expressing Cry1Ac protein as compared to Cry1Ab to Helicoverpa armigera. Pyramided transgenic plants with moderate expression levels (15–20 ng mg−1 of TSP) showed high-level of resistance and protection against pod borer larvae of H. armigera as compared to high level expression of a single toxin. These results have shown the significance of pyramiding and co-expression of two Cry toxins for efficient protection against lepidopteran pests of chickpea.
http://www.springerlink.com/content/g2978h5u4123t276/
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