Article

Expression of δ-endotoxin Cry1EC from an inducible promoter confers insect protection in peanut (Arachis hypogaea L.) plants.

National Agri-Food Biotechnology Institute, Department of Biotechnology, Govt of India, Mohali, Punjab, India.
Pest Management Science (Impact Factor: 2.59). 10/2010; 67(2):137-45. DOI: 10.1002/ps.2041
Source: PubMed

ABSTRACT Spodoptera litura (F.) is a polyphagous foliage insect and a major pest on peanut (Arachis hypogaea L.). Constitutive expression of δ-endotoxin Cry1EC gives protection against S. litura, as reported earlier. In this study, insect bites and salicylic acid induced high-level expression of Cry1EC was achieved in peanut. In order to achieve this, the expression of pathogenesis responsive promoter PR-1a was enhanced by placing it downstream of the CaMV35S promoter in the pCAMBIA 1300 backbone. The resultant promoter CaMV35S(r)PR-1a expressed a high level of insecticidal δ-endotoxin Cry1EC. The Gus expression under the control of CaMV35S(r)PR-1a served as a convenient marker for evaluation of promoter response to different treatments.
Transgenic events that showed a very low level of uninduced expression and no expression in seeds were selected. The Cry1EC expression in leaves increased nearly eightfold in the selected event, following induction by salicylic acid. Both the salicylic-acid-treated and the S. litura-bitten leaves showed the highest expression after 2 days. Leaves from salicylic-acid-induced transgenic plants caused 100% mortality of S. litura at all stages of larval development.
The results suggest that high expression of inducible promoters provides a good strategy for the development of safer transgenic food and feed crops.

0 Bookmarks
 · 
90 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: De-embryonated cotyledon explants of peanut were co-cultivated under different conditions with Agrobacterium tumefaciens harbouring pIG121hm plasmid carrying intron-containing β-glucuronidase as a reporter while hygromycin phosphotransferase and neomycin phosphotransferase as selectable marker genes. Co-cultivation duration and temperature, various antioxidants and their concentrations, bacterial strains and explant characteristics (incised and non-incised) were examined either alone or in combinations for optimization of transient expression of the reporter gene. Up to 81% transformation was recorded when non-incised explants were co-cultivated with strain EHA101 for 5 days at 21°C on shoot induction medium containing 100 mg/L l-cysteine. Addition of the optimized concentration of augmentin (200 mg/L) along with cefotaxime (200 mg/L) to the shoot induction medium not only effectively eliminated bacterial growth, but also facilitated high frequency of shoot induction. The 40 mg/L hygromycin concentration prevented complete shoot regeneration of non-transgenic explants thus considered for the regeneration of transgenics. Resistant shoots were successfully transferred to soil either by grafting or in vitro rooting. Survival rate of the grafted shoots was nearly 100% in glass-house conditions. The optimized protocol took around 3 months to generate healthy plants. Polymerase chain reaction, Southern blot hybridization, histochemical tests, segregation and hygromycin-leaf assays of selected transgenic plants showed integration of the transgene into peanut genome. No chimeras were noticed during the study.
    Plant Cell Tissue and Organ Culture 109(1). · 3.63 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: KEY MESSAGE : Peanuts transformed with the synthetic cry8Ea1 gene flanked by MARs are a potentially effective control strategy against white grubs. Cry8Ea1 protein levels of the construct containing MARs were increased by 2.5 times. White grubs are now recognized as the most important pests of peanut worldwide. A synthetic cry8Ea1 gene, which was toxic to Holotrichia parallela larvae, was expressed in chimeric peanut roots using an Agrobacterium rhizogenes-mediated transformation system. The relative mRNA and protein levels of the cry8Ea1 gene were confirmed by quantitative real-time PCR and ELISA, respectively. The effects of matrix attachment regions (MARs) on the expression and activity of the cry8Ea1 gene were analyzed. The average expression level of cry8Ea1 in peanut roots was higher for the plants harboring constructs flanked by MARs from tobacco. Moreover, differing from previous studies, the synthetic cry8Ea1 gene flanked by MARs showed more variation in protein levels than mRNA levels. These composite plants containing cry8Ea1 gene flanked by MARs exhibited a high toxicity against Holotrichia parallela larvae as shown by bioassay analysis, thus offering a potential effective combination to control subterranean insects in peanuts.
    Plant Cell Reports 03/2013; · 2.51 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Aphid Aphid s are economically significant agricultural pests that are responsible for large yield losses in many different crops. Because the use of insecticides is restricted in the context of integrated pest management and aphids develop resistance against them rapidly, new biotechnology-based approaches are required for aphid control. These approaches focus on the development of genetically modified aphid-resistant plants that express protease inhibitor Protease inhibitor s, dsRNA, antimicrobial peptide antimicrobial peptide s, or repellent Repellent s, thus addressing different levels of aphid-plant interactions. However, a common goal is to disturb host plant acceptance by aphid Aphid s and to disrupt their ability to take nutrition from plants. The defense agents negatively affect different fitness-associated parameters such as growth, reproduction, and survival, which therefore reduce the impact of infestations. The results from several different studies suggest that biotechnology-based approaches offer a promising strategy for aphid control.
    Advances in biochemical engineering/biotechnology 06/2013; · 1.64 Impact Factor

Full-text

View
107 Downloads
Available from
Jun 5, 2014