Transgenic rice expressing Allium sativum leaf lectin with enhanced resistance against sap-sucking insect pests |
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Authors: | Prasenjit Saha Pralay Majumder Indrajit Dutta Tui Ray S C Roy Sampa Das |
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Institution: | (1) Plant Molecular and Cellular Genetics, Bose Institute, P1/12 C.I.T. Scheme VII (M), 700054 Kolkata, India;(2) Centre of Advanced Study, Cell and Chromosome Research, Department of Botany, University of Calcutta, 35 Ballygunge Circular Road, 700019 Kolkata, India |
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Abstract: | Mannose binding Allium sativum leaf agglutinin (ASAL) has been shown to be antifeedant and insecticidal against sap-sucking insects. In the present investigation, ASAL coding sequence was expressed under the control of CaMV35S promoter in a chimeric gene cassette containing plant selection marker, hpt and gusA reporter gene of pCAMBIA1301 binary vector in an elite indica rice cv. IR64. Many fertile transgenic plants were generated using scutellar calli as initial explants through Agrobacterium-mediated transformation technology. GUS activity was observed in selected calli and in mature plants. Transformation frequency was calculated to be ~12.1%±0.351 (mean ± SE). Southern blot analyses revealed the integration of ASAL gene into rice genome with a predominant single copy insertion. Transgene localization was detected on chromosomes of transformed plants using PRINS and C-PRINS techniques. Northern and western blot analyses determined the expression of transgene in transformed lines. ELISA analyses estimated ASAL expression up to 0.72 and 0.67% of total soluble protein in T0 and T1 plants, respectively. Survival and fecundity of brown planthopper and green leafhopper were reduced to 36% (P<0.01), 32% (P<0.05) and 40.5, 29.5% (P<0.001), respectively, when tested on selected plants in comparison to control plants. Specific binding of expressed ASAL to receptor proteins of insect gut was analysed. Analysis of T1 progenies confirmed the inheritance of the transgenes. Thus, ASAL promises to be a potential component in insect resistance rice breeding programme. |
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Keywords: | Allium sativum leaf lectin Brown planthopper Cycling-primed in situ labelling Green leafhopper Insect bioassay Rice cv IR64 |
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