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Agrobacterium mediated transformation of indica rice (Oryza sativa L.) for insect resistance

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Abstract

The rice leaffolder (RLF), Cnaphalocrocis medinalis is an important pest of rice that causes severe damage in many areas of the world. The plants were transformed with fully modified (plant codon optimized) synthetic Cry1C coding sequences as well as with the hpt and gus genes, coding for hygromycin phosphotransferase and β-glucuronidase, respectively. Cry1C sequences placed under the control of doubled 35S promoter plus the AMV leader sequence, and hpt and gus genes driven by cauliflower mosaic virus 35S promoter, were used in this study. Embryogenic calli after cocultivation with Agrobacterium were selected on the medium containing hygromycin B. A total of 67 hygromycin-resistant plants were regenerated. PCR and Southern blot analyses of primary transformants revealed the stable integration of Cry1C coding sequences into the rice genome with predominant single copy integration. R1 progeny plants disclosed a monogenic pattern (3:1) of transgene segregation as confirmed by molecular analyses. These transgenic lines were highly resistant to rice leaffolder (RLF), Cnaphalocrocis medinalis as revealed by insect bioassay.

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References

  • Alcantara EP, Aguda RM, Curtiss A, Dean DH, Cohen MB (2004) Bacillus thuringiensis δ-endotoxin binding to brush border membrane vesicles of rice stem borers. Arch Insect Biochem Physiol 55:169–177

    Article  PubMed  CAS  Google Scholar 

  • Aragao FJL, Barros LMG, de Sousa MV, de Sa Grossi MF, Almeida ERP, Gander ES, Rech EL (1999) Expression of a methionine-rich storage albumin from the brazil nut (Bertholletia excelsa H.B.K., Lecythidaceae) in transgenic bean plants (Phaseolus vulgaris L., Fabaceae). Genet Mol Biol 22:445–449

    Article  CAS  Google Scholar 

  • Bashir K, Husnain T, Fatima T, Riaz N, Makhdoom R, Riazuddin S (2005) Novel indica basmati line (B-370) expressing two unrelated genes of Bacillus thuringiensis is highly resistant to two lepidopteran insects in the field. Crop Prot 24:870–879

    Article  CAS  Google Scholar 

  • Bower R, Elliott AR, Potier BAM, Birch RG (1996) High-efficiency, microprojectile mediated cotransformation of sugarcane, using visible or selectable markers. Mol Breed 2:239–249

    Article  CAS  Google Scholar 

  • Brookes P, Barfoot GB (2003) GM rice, will this be the way for global acceptance of GM crop technology. ISAAA Briefs no. 28, ISAAA, Ithaca

  • Cao J, Tang JD, Strizhov N, Shelton AM, Earle ED (1999) Transgenic broccoli with high levels of Bacillus thuringiensis Cry1C protein control diamondback moth larvae resistant to Cry1A or Cry1C. Mol Breed 5:131–141

    Article  CAS  Google Scholar 

  • Chen M, Zhao JZ, Ye GY, Fu Q, Shelton AM (2006) Impact of insect-resistant transgenic rice on target insect pests and non-target arthropods in China. Insect Sci 13:409–420

    CAS  Google Scholar 

  • Cheng X, Sardana R, Kaplan H, Altosaar I (1998) Agrobacterium transformed rice plants expressing synthetic Cry IA(b) and Cry IA(c) genes are highly toxic to stripe and stem borer and yellow stem borer. Proc Natl Acad Sci USA 95:2767–2772

    Article  PubMed  CAS  Google Scholar 

  • Chilton MD, Currier TC, Farrand SK, Bendich AJ, Gordon MP, Nester EW (1974) Agrobacterium tumefaciens DNA and PS8 bacteriophage DNA not detected in crown gall tumours. Proc Nat Acad Sci USA 71:3672–3676

    Article  PubMed  CAS  Google Scholar 

  • Christou P, Ford TL, Kofron M (1992) The development of a variety independent gene transfer method for rice. Tibtech 10:239–246

    Google Scholar 

  • Dale D (1994) Insect pests of the rice plant–their biology and ecology. In: Heinrichs EA (ed) Biology and management of rice insects. Wiley Eastern/New Age International, New Delhi, India, pp 363–485

    Google Scholar 

  • Desneux N, Decourtye A, Delpuech JM (2007) The sublethal effects of pesticides on beneficial arthropods. Annu Rev Entomol 52:81–106

    Article  PubMed  CAS  Google Scholar 

  • FAO (2007) FAOSTAT. http://faostat.fao.org/site/567/default.aspx

  • Ferry N, Edwards MG, Gatehouse J, Capell T, Christou P, Gatehouse AMR (2006) Transgenic plants for insect pest control: a forward looking scientific perspective. Transgenic Res 15:13–19

    Article  PubMed  CAS  Google Scholar 

  • Fujimoto H, Itoh K, Yamamoto M, Kyozuka J, Shimamoto K (1993) Insect resistant rice generated by introduction of a modified dendotoxin gene of Bacillus thuringiensis. Bio/Technol 11:1151–1155

    CAS  Google Scholar 

  • Gatehouse JA (2008) Biotechnological prospects for engineering insect-resistant plants. Plant Physiol 146:881–887

    Article  PubMed  CAS  Google Scholar 

  • He Y, Jones HD, Chen S, Chen XM, Wang DW, Li KX, Wang DS, Xia LQ (2010) Agrobacterium-mediated transformation of durum wheat (Triticum turgidum L. var. durum cv. Stewart) with improved efficiency. J Exp Bot 61:1567–1581

    Article  PubMed  CAS  Google Scholar 

  • Hiei Y, Komari T (2006) Improved protocols for transformation of indica rice mediated by Agrobacterium tumefaciens. Plant Cell Tiss Organ Cult 85:271–283

    Article  CAS  Google Scholar 

  • Hiei Y, Ohta S, Komari T, Kumashiro T (1994) Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J 6:271–282

    Article  PubMed  CAS  Google Scholar 

  • Holsters M, de Waele D, Depicker A, Messens E, van Montagu M, Schell J (1978) Transfection and transformation of Agrobacterium tumefaciens. Mol Gen Genet 163:181–187

    Article  PubMed  CAS  Google Scholar 

  • James C (2005) Global status of commercialized biotech/GM crops: 2005. ISAAA brief no. 34. International Service for the Acquisition of Agri-Biotech Applications, Ithaca, New York, USA

    Google Scholar 

  • Jefferson RA (1987) Assaying chimeric genes in plants: the GUS gene fusion system. Plant Mol Biol Rep 5:387–405

    Article  CAS  Google Scholar 

  • Khan SA, Zafar Y, Briddon RW, Malik KA, Mukhtar Z (2006) Spider venom toxin protects plants from insect attack. Transgenic Res 15:349–357

    Article  PubMed  CAS  Google Scholar 

  • Khanna HK, Raina SK (1999) Agrobacterium-mediated transformation of indica rice cultivars using binary and superbinary vectors. Aust J Plant Physiol 26:311–324

    Article  CAS  Google Scholar 

  • Khush G (1997) Origin, dispersal, cultivation and variation in rice. Plant Mol Biol 35:25–34

    Article  PubMed  CAS  Google Scholar 

  • Maqbool SB, Raizuddin S, Loc TN, Gatehouse AMR, Gatehouse JA, Christou P (2001) Expression of multiple insecticidal genes confers broad resistance against a range of different rice pests. Mol Breed 7:85–93

    Article  CAS  Google Scholar 

  • Matteson PC (2000) Insect pest management in tropical Asian irrigated rice. Annu Rev Entomol 45:549–574

    Article  PubMed  CAS  Google Scholar 

  • Mazier M, Pennetier C, Tourneur J, Jounin L, Giband M (1997) The expression of Bacillus thuringiensis toxin genes in plant cells. Biotechnol Annu Rev 3:313–347

    Article  CAS  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  • Nayak P, Basu D, Das S, Basu A, Ghosh D, Ramakrishnan NA, Ghosh M, Sen SK (1997) Transgenic elite indica rice plants expressing Cry IAc d-endotoxin of Bacillus thuringiensis are resistant against yellow stem borer (Scirpophaga incertulas). Proc Natl Acad Sci USA 94:2111–2116

    Article  PubMed  CAS  Google Scholar 

  • Qiu HJ, Wei ZM, An HL, Zhu YX (2001) Agrobacterium tumefaciens-mediated transformation of rice with the spider insecticidal gene conferring resistance to leaf folder and striped stem borer. Cell Res 11:149–155

    Google Scholar 

  • Rahman M, Rashid H, Shahid AA, Bashir K, Husnain T, Riazuddin S (2007) Insect resistance and risk assessment studies of advanced generations of basmati rice expressing two genes of Bacillus thuringiensis. Plant Biotech. doi:10.2225/vol10-issue2-fulltext-3

  • Ramesh S, Nagadhara D, Reddy VD, Rao KV (2004) Production of transgenic indica rice resistant to yellow stem borer and sap-sucking insects, using super-binary vectors of Agrobacterium tumefaciens. Plant Sci 166:1077–1085

    Article  CAS  Google Scholar 

  • Rashid H, Yokoi S, Toriyama K, Hinata K (1996) Transgenic plant production mediated by Agrobacterium in indica rice. Plant Cell Rep 15:727–730

    Article  CAS  Google Scholar 

  • Raveendar S, Premkumar A, Ignacimuthu S, Agastian P (2008) Effect of sea water on callus induction and regeneration of rice genotypes. Int J Integr Biol 3:92–95

    CAS  Google Scholar 

  • Romeis J, Michael M, Franz B (2006) Transgenic crops expressing Bacillus thuringiensis toxins and biological control. Nat Biotechnol 24:63–71

    Article  PubMed  CAS  Google Scholar 

  • Savary S, Willocquet L, Elazegui FS, Castilla NP, Teng PS (2000) Rice pest constraints in tropical Asia: quantification of yield losses due to rice pests in a range of production situations. Plant Dis 84:357–369

    Article  Google Scholar 

  • Shu QY, Ye GY, Cui HR, Cheng XY, Xiang YB, Wu DX, Gao MW, Xia YW, Hu C, Sardana R, Altosaar I (2000) Transgenic rice plants with a synthetic cry1Ab gene from Bacillus thuringiensis were highly resistant to eight lepidopteran rice pest species. Mole Breed 6:433–439

    Article  CAS  Google Scholar 

  • Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517

    Article  PubMed  CAS  Google Scholar 

  • Strizhov N, Keller M, Mathur J (1996) A synthetic cry IC gene, encoding a Bacillus thuringiensis d endotoxin, confers Spodoptera resistance in alfalfa and tobacco. Proc Natl Acad Sci USA 93:15012–15017

    Article  PubMed  CAS  Google Scholar 

  • Tang W, Chen H, Xu CG, Li XH, Lin YJ, Zhang QF (2006) Development of insect-resistant transgenic indica rice with a synthetic cry1C* gene. Mole Breed 18:1–10

    Article  CAS  Google Scholar 

  • Tu J, Zhang G, Datta K, Xu C, He Y, Zhang Q, Khush GS, Datta SK (2000) Field performance of transgenic elite commercial hybrid rice expressing Bacillus thuringiensis d-endotoxin. Nat Biotechnol 18:1101–1104

    Article  PubMed  CAS  Google Scholar 

  • Wilmink A, Dons JM (1993) Selective agents and marker genes for use in transformation of monocotyledonous plants. Plant Mol Biol Rep 11:165–185

    Article  CAS  Google Scholar 

  • Xia H, Chen L, Wang F, Lu BR (2010) Yield benefit and underlying cost of insect-resistance transgenic rice: implication in breeding and deploying transgenic crops. Field Crop Res 118:215–220

    Article  Google Scholar 

  • Xiaofen S, Kexuan T, Bingliang W, Huaxiong Q, Xinggu L (2001) Transgenic rice homozygous lines expressing GNA showed enhanced resistance to rice brown planthopper. Chin Sci Bull 46:1698–1703

    Article  Google Scholar 

  • Ye GY, Tu J, Hu C, Datta K, Datta SK (2001) Transgenic IR72 with fused Bt gene cry1Ab/cry1Ac from Bacillus thuringiensis is resistant against four lepidopteran species under field conditions. Plant Biotechnol 18:125–133

    Article  CAS  Google Scholar 

  • Ye GY, Yao HW, Shu QY, Cheng XY, Hu C, Xia YW, Gao MW, Altosaar I (2003) High levels of stable resistance in transgenic rice with a cry1Ab gene from Bacillus thuringiensis Berliner to rice leaf folder, Cnaphalocrocis medinalis (Guenee) under field conditions. Crop Prot 22:171–178

    Article  CAS  Google Scholar 

  • Ye R, Huang H, Yang Z, Chen T, Liu L, Li X, Chen H, Lin Y (2009a) Development of insect-resistant transgenic rice with Cry1C free endosperm. Pest Manag Sci 65:1015–1020

    Article  PubMed  CAS  Google Scholar 

  • Ye SH, Chen S, Zhang F, Wang W, Tian Q, Liu JZ, Chen F, Bao JK (2009b) Transgenic tobacco expressing Zephyranthes grandiflora agglutinin confers enhanced resistance to aphids. Appl Biochem Biotechnol 158:615–630

    Article  PubMed  CAS  Google Scholar 

  • Zaidi MA, Ye G, Yao H, You TH, Loit E, Dean DH, Riazuddin S, Altosaar I (2009) Transgenic rice plants expressing a modified cry1Ca1 gene are resistant to Spodoptera litura and Chilo suppressalis. Mol Biotechnol 43:232–242

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

We thank Prof. Dr. Illimar Altosar, University of Ottawa, Canada for providing the binary vector and Dr. Gabrial Paulraj, Scientist and Mr K. Baskar, Entomology Research Institute, Loyola College, Chennai for their help and support during insect bioassay. We are grateful to Entomology Research Institute for financial assistance.

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  1. Plant Biotechnology Unit, Entomology Research Institute, Loyola College, Chennai, 600 034, Tamil Nadu, India

    S. Ignacimuthu & S. Raveendar

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  1. S. Ignacimuthu
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  2. S. Raveendar
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Correspondence to S. Ignacimuthu.

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Ignacimuthu, S., Raveendar, S. Agrobacterium mediated transformation of indica rice (Oryza sativa L.) for insect resistance. Euphytica 179, 277–286 (2011). https://doi.org/10.1007/s10681-010-0308-7

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  • DOI: https://doi.org/10.1007/s10681-010-0308-7

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