, Volume 195, Issue 1, pp 129–142 | Cite as

Development of elite rice restorer lines in the genetic background of R022 possessing tolerance to brown planthopper, stem borer, leaf folder and herbicide through marker-assisted breeding

  • Bingliang Wan
  • Zhongping Zha
  • Jinbo Li
  • Mingyuan Xia
  • Xueshu Du
  • Yongjun Lin
  • Desuo Yin


Rice leaf folder, stem borer and brown planthopper (BPH) are the most devastating rice insect pests. Developing and planting insect-resistant rice varieties is the most economical and effective measure for controlling these pests. BPH can be controlled with native BPH-resistance genes in rice, while at present rice leaf folder and stem borer can only be controlled through planting transgenic Bt rice. In this study, the breeding of a new restorer line KR022 possessing stacked BPH-resistance genes Bph14 and Bph15, Bt gene cry1C and glufosinate-resistance gene bar, is reported for the first time. A rice restorer line R022 with BPH-resistance genes Bph14 and Bph15 was used as a recurrent parent to cross with the transgenic rice T1C-19 of cry1C and bar genes during the breeding process. The restorer line KR022 was developed from the backcross populations of R022 and T1C-19 through molecular marker-assisted selection and glufosinate-resistance selection. The cry1C and bar genes were found to integrate on chromosome 11 of KR022, and the genome recovery of KR022 was up to 95.8 % of the R022 genome. The quantification of Cry1C protein expression showed that it was expressed at different levels in the leaf, stem, panicle, endosperm, and root of KR022 and its hybrid rice. The insect-resistance evaluation indicated that KR022 and its hybrid rice had good resistance to rice leaf folder and stem borer, both in laboratory settings and in the field. Furthermore, they exhibited increased resistance to BPH at both the seedling and mature stage. The field trial showed there was no significant difference in key agronomic traits between KR022 and its recurrent parent R022, and four hybrids from KR022 yield much higher than the control II-You 838. Moreover, KR022 and its hybrid rice were found to have resistance to the herbicide glufosinate. These results demonstrate that KR022 is effective as a rice restorer line for the breeding of “green super rice”, possessing multiple tolerances to rice BPH, stem borer, leaf folder and glufosinate.


bar Bph14/Bph15 cry1C Leaf folder resistance Stem borer resistance Oryza sativa L. 



Brown planthopper


Bacillus thuringiensis


Deoxy-ribonucleoside triphosphate


Enzyme-linked immunosorbent assay




Molecular marker-assisted selection


Polymerase chain reaction




Striped stem borer


Yellow stem borer



We thank Dr. Weihua Ma in the National Key Laboratory of Crop Genetic Improvement and National Center of Plant Gene Research (Wuhan, China) for the help in SSB bioassay. This work was supported by the National Program of Transgenic Variety Development of China (2011ZX08001-001) and the Hubei Centre of Innovation for Agricultural Science and Technology (China).


  1. Bharathkumara S, Paulraja RSD, Brindhaa PV, Kavithaa S, Gnanamanickam SS (2008) Improvement of bacterial blight resistance in rice cultivars Jyothi and IR50 via marker-assisted backcross breeding. J Crop Improve 21:101–116CrossRefGoogle Scholar
  2. Block MD, Botterman J, Vandewiele M, Dockx J, Thoen C, Gosselé V, Movva NR, Thompson C, Montagu MV, Leemans J (1987) Engineering herbicide resistance in plants by expression of a detoxifying enzyme. EMBO J 6:2513–2518PubMedGoogle Scholar
  3. Block MD, Brouwer DD, Tenning P (1989) Transformation of Brassica napus and Brassica oleracea using Agrobacterium tumefaciens and the expression of the bar and neo genes in the transgenic plants. Plant Physol 91:694–701CrossRefGoogle Scholar
  4. Chen XW, Li SG, Ma YQ, Li HY, Zhou KD, Zhu LH (2004) Marker-assisted selection and pyramiding for three blast resistance genes, Pi-d(t)1, Pi-b, Pi-ta2, in rice. Chin J Biotech 20:708–714Google Scholar
  5. Chen H, Tang W, Xu C, Li X, Lin Y, Zhang Q (2005) Transgenic indica rice plants harboring a synthetic cry2A* gene of Bacillus thuringiensis exhibit enhanced resistance against lepidopteran rice pests. Theor Appl Genet 111:1330–1337PubMedCrossRefGoogle Scholar
  6. Cornejo MJ, Luth D, Blankenship KM, Anderson OD, Blechl AE (1993) Activity of a maize ubiquitin promoter in transgenic rice. Plant Mol Biol 23:567–581PubMedCrossRefGoogle Scholar
  7. Datta K, Baisakh N, Maung Thet K, Tu J, Datta SK (2002) Pyramiding transgenes for multiple resistance in rice against bacterial blight, yellow stem borer and sheath blight. Theor Appl Genet 106:1–8PubMedGoogle Scholar
  8. Du B, Zhang W, Liu B, Hu J, Wei Z, Shi Z, He R, Zhu L, Chen R, Han B, He G (2009) Identification and characterization of Bph14, a gene conferring resistance to brown planthopper in rice. Proc Natl Acad Sci USA 106:22163–22168PubMedCrossRefGoogle Scholar
  9. Farooq M, Siddique KHM, Rehman H, Aziz T, Leed DJ, Wahide A (2011) Rice direct seeding: experiences, challenges and opportunities. Soil Tillage Res 111:87–98CrossRefGoogle Scholar
  10. Fu YP, Zhu ZG, Xiao H, Hu GC, Si HM, Yu YH, Sun ZX (2001) Primary study of mechanization of seed production of hybrid rice by introducing bar gene to Pei’ai 64S. Chinese J Rice Sci 15:97–100Google Scholar
  11. Fujimoto H, Itoh K, Yamamoto M, Kyozuka J, Shimamoto K (1993) Insect resistant rice generated by introduction of a modified δ-endotoxin gene of Bacillus thuringiensis. Nat Biotech 11:1151–1155CrossRefGoogle Scholar
  12. Gressel J, Valverde BE (2009) A strategy to provide long-term control of weedy rice while mitigating herbicide resistance transgene flow, and its potential use for other crops with related weeds. Pest Manag Sci 65:723–731PubMedCrossRefGoogle Scholar
  13. Harini AS, Lakshmi SS, Kumar SS, Sivaramakrishnan S, Kadirvel P (2010) Validation and fine-mapping of genetic locus associated with resistance to brown plant hopper [Nilaparvata lugens (Stal.)] in rice (Oryza sativa L.). Asian J Bio Sci 5:32–37Google Scholar
  14. Hu J, Li X, Wu C, Yang C, Hua H, Gao G, Xiao J, He Y (2012) Pyramiding and evaluation of the brown planthopper resistance genes Bph14 and Bph15 in hybrid rice. Mol Breed 29:61–69CrossRefGoogle Scholar
  15. Huang D, Li J, Zhang S, Xue R, Yang W, Hua Z, Xie X, Wang X (1998) New technology to examine and improve the purity of hybrid rice with herbicide resistant gene. Chin Sci Bull 43:784–787CrossRefGoogle Scholar
  16. Huang J, Hu R, Pray C, Qiao F, Rozelle S (2003) Biotechnology as an alternative to chemical pesticides: a case study of Bt cotton in China. Agri Econ 29:55–67CrossRefGoogle Scholar
  17. Hyde J, Martin MA, Preckel PV, Edwards CR (1999) The economics of Bt corn: valuing protection from the European corn borer. Appl Econ Perspect Pol 21:442–454CrossRefGoogle Scholar
  18. Jena KK, Kim SM (2010) Current status of brown planthopper (BPH) resistance and genetics. Rice 3:161–171CrossRefGoogle Scholar
  19. Jiang GH, Xu CG, Tu JM, Li XH, He YQ (2004) Pyramiding of insect-and disease-resistance genes into an elite indica, cytoplasm male sterile restorer line of rice, ‘Minghui 63′. Plant Breed 123:112–116CrossRefGoogle Scholar
  20. Li J, Xia M, Qi H, He G, Wan B, Zha Z (2006) Marker-assisted selection for brown planthopper (Nilaparvata lugens Stl) resistance genes Bph14 and Bph15 in rice. Sci Agric Sin 39:2132–2137Google Scholar
  21. Li J, Wan B, Xia M, Qi H, Shi H, Xin F (2011) Breeding of the brown planthopper resistant rice varieties. Chin J Appl Entomol 48:1348–1353Google Scholar
  22. Lu L, Wu X, Yin X, Morrand J, Chen X, Folk WR, Zhang Z (2009) Development of marker-free transgenic sorghum [Sorghum bicolor (L.) Moench] using standard binary vectors with bar as a selectable marker. Plant Cell Tiss Organ Cult 99:97–108CrossRefGoogle Scholar
  23. Myint KKM, Fujita D, Matsumura M, Sonoda T, Yoshimura A, Yasui H (2012) Mapping and pyramiding of two major genes for resistance to the brown planthopper (Nilaparvata lugens [Stål]) in the rice cultivar ADR52. Theor Appl Genet 124:495–504PubMedCrossRefGoogle Scholar
  24. Peñalver Cruz A, Arida A, Heong KL, Horgan FG (2011) Aspects of brown planthopper adaptation to resistant rice varieties with the Bph3 gene. Entomol Exp Appl 141:245–257CrossRefGoogle Scholar
  25. Phipps RH, Park JR (2002) Environmental benefits of genetically modified crops: global and European perspectives on their ability to reduce pesticide use. J Anim Feed Sci 11:1–18Google Scholar
  26. Pray CE, Jhuang J, Hu R, Rozelle S (2002) Five years of Bt cotton in China the benefits continue. Plant J 31:423–430PubMedCrossRefGoogle Scholar
  27. Rooke L, Byrne D, Salgueiro S (2005) Marker gene expression driven by the maize ubiquitin promoter in transgenic wheat. Ann Appl Biol 136:167–172CrossRefGoogle Scholar
  28. Song X, Liu L, Wang Z, Qiang S (2009) Potential gene flow from transgenic rice (Oryza sativa L.) to different weedy rice (Oryza sativa f. spontanea) accessions based on reproductive compatibility. Pest Manag Sci 65:862–869PubMedCrossRefGoogle Scholar
  29. Stewart SD, Adamczyk JJ, Knighten KS, Davis FM (2001) Impact of Bt cottons expressing one or two insecticidal proteins of Bacillus thuringiensis Berliner on growth and survival of noctuid (Lepidoptera) larvae. J Econ Entomol 94:752–760PubMedCrossRefGoogle Scholar
  30. Tang W, Chen H, Xu C, Li X, Lin Y, Zhang Q (2006) Development of insect-resistant transgenic indica rice with a synthetic cry1C* gene. Mol Breed 18:1–10CrossRefGoogle Scholar
  31. Toki S, Takamatsu S, Nojiri C, Ooba S, Anzai H, Iwata M, Christensen AH, Quail PH, Uchimiya H (1992) Expression of a maize ubiquitin gene promoter-bar chimeric gene in transgenic rice plants. Plant Physol 100:1503–1507CrossRefGoogle Scholar
  32. 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 δ-endotoxin. Nat Biotech 18:1101–1104CrossRefGoogle Scholar
  33. Wang Y (2006) Analysis on the occurrence and development of rice diseases and insects in China. Chin Agric Sci Bull 22:343–347Google Scholar
  34. Wang ZX, Wang XJ, Jia SR (2011) Data survey and analysis of the gene flow frequencies and distances in major crops II. Rice J Agri Sci Technol 13:30–34Google Scholar
  35. Wei Y, Yao F, Zhu C, Jiang M, Li G, Song Y, Wen F (2008) Breeding of transgenic rice restorer line for multiple resistance against bacterial blight, striped stem borer and herbicide. Euphytica 163:177–184CrossRefGoogle Scholar
  36. Wu M, Lin J, Hua Z, Yan M, Huang D (2000) Application of bar gene in directed seeded rice. Acta Agriculturae Zhejiangensis 12:290–293Google Scholar
  37. Yang H, You A, Yang Z, Zhang F, He R, Zhu L, He G (2004) High-resolution genetic mapping at the Bph15 locus for brown planthopper resistance in rice (Oryza sativa L.). Theor Appl Genet 10:182–191CrossRefGoogle Scholar
  38. Yang Z, Chen H, Tang W, Hua H, Lin Y (2011) Development and characterization of transgenic rice expressing two Bacillus thuringiensis genes. Pest Manag Sci 67:414–422PubMedCrossRefGoogle Scholar
  39. Yao KM, Hu N, Chen WL, Li RZ, Yuan QH, Wang F, Qian Q, Jia SR (2008) Establishment of a rice transgene flow model for predicting maximum distances of gene flow in southern China. New Phyt 180:217–228CrossRefGoogle Scholar
  40. Ye G, 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 18:125–133CrossRefGoogle Scholar
  41. Yorobe JM, Quicoy CB (2006) Economic impact of Bt corn in the Philippines. Philipp Agric Sci 89:258–267Google Scholar
  42. Zhang Q (2007) Strategies for developing green super rice. Proc Natl Acad Sci USA 104:16402–16409PubMedCrossRefGoogle Scholar
  43. Zheng X, Yang Y, Xu H, Chen H, Wang B, Lin Y, Lu Z (2011) Resistance performances of transgenic Bt rice lines T2A-1 and T1c-19 against Cnaphalocrocis medinalis (Lepidoptera: pyralidae). J Econom Entomol 104:1730–1735CrossRefGoogle Scholar
  44. Zhou Y, Xu J, Zhou S, Yu J, Xie X, Xu M, Sun Y, Zhu L, Fu B, Gao Y (2009) Pyramiding Xa23 and Rxo1 for resistance to two bacterial diseases into an elite indica rice variety using molecular approaches. Mol Breed 23:279–287CrossRefGoogle Scholar
  45. Zuo J, Zhang LJ, Song XL, Dai WM, Qiang S (2011) Innate factors causing differences in gene flow frequency fromtransgenic rice to different weedy rice biotypes. Pest Manag Sci 67:677–690PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Bingliang Wan
    • 1
  • Zhongping Zha
    • 1
  • Jinbo Li
    • 1
  • Mingyuan Xia
    • 1
  • Xueshu Du
    • 1
  • Yongjun Lin
    • 2
  • Desuo Yin
    • 1
  1. 1.Hubei Key Laboratory of Food Crop Germplasm and Genetic Improvement, Food Crop Research InstituteHubei Academy of Agricultural ScienceWuhanPeople’s Republic of China
  2. 2.National Key Laboratory of Crop Genetic Improvement and National Centre of Plant Gene ResearchHuazhong Agricultural UniversityWuhanPeople’s Republic of China

Personalised recommendations