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Theoretical and Applied Genetics

, Volume 91, Issue 1, pp 9–14 | Cite as

Development of a PCR-based marker to identify rice blast resistance gene, Pi-2(t), in a segregating population

  • S. Hittalmani
  • M. R. Foolad
  • T. Mew
  • R. L. Rodriguez
  • N. Huang
Article

Abstract

The genomic clone RG64, which is tightly linked to the blast resistance gene Pi-2(t) in rice, provides means to perform marker-aided selection in a rice breeding program. The objective of this study was to investigate the possibility of generating a polymerase chain reaction (PCR)-based polymorphic marker that can distinguish the blast resistant gene, Pi-2(t), and susceptible genotypes within cultivated rice. RG64 was sequenced, and the sequence data was used to design pairs of specific primers for (PCR) amplification of genomic DNA from rice varieties differing in their blast disease responsiveness. The amplified products, known as sequenced-tagged-sites (STSs), were not polymorphic between the three varieties examined. However, cleavage of the amplified products with the restriction enzyme HaeIII generated a polymorphic fragment, known as specific amplicon polymorphism (SAP), between the resistant and the susceptible genotypes. To examine the power of the identified SAP marker in predicting the genotype of the Pi-2 (t) locus, we determined the genotypes of the F2 individuals at this locus by performing progeny testing for the disease response in the F3 generation. The results indicated an accuracy of more than 95% in identifying the resistant plants, which was similar to that using RG64 as the hybridization probe. The identification of the resistant homozygous plants increased to 100% when the markers flanking the genes were considered simultaneously. These results demonstrate the utility of SAP markers as simple and yet reliable landmarks for use in marker-assisted selection and breeding within cultivated rice.

Key words

Molecular markers RFLP PCR SAPs DNA sequence Marker-assisted selection 

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References

  1. Abenes MLP, Angeles EP, Khush GS, Huang N (1993) Selection of bacterial blight resistant plants in F2 generation via their linkage to molecular markers. Rice Genet Newsl 10:120–123Google Scholar
  2. Biggin MD, Gibson TJ, Hong GF (1993) Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci USA 80:3963–3965Google Scholar
  3. Bonman JM, Mackill DJ (1988) Durable resistance to rice blast disease. Oryza 25:103–110Google Scholar
  4. Bureau ET, Wessler SR (1994) Mobile inverted repeat elements of the Tourist family are associated with the genes of many cereal grasses. Proc Natl Acad Sci USA 91:1411–1415Google Scholar
  5. Chunwongse J, Martin GB, Tanksley SD (1992) Pre-germination, genotypic screening using PCR amplification on half seeds. Theor Appl Genet 86:694–698Google Scholar
  6. Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation: version II. Plant Mol Biol Rep 1:19–21Google Scholar
  7. Henikoff S (1988) Unidirectional digestion with exonuclease III creates targeted breakpoints for DNA sequencing. Gene 28:351–359Google Scholar
  8. Kleinhofs A, Kilian A, Saghai Maroof MA, Biyashev RM, Hayes P, Chen FQ, Lapitan N, Fenwick A, Blake TK, Kanazin V, Ananiev E, Dahleen L, Kudrna D, Bollinger J, Knapp SJ, Liu B, Sorrels M, Heun M, Franckowiak JD, Hoffman D, Skadsen R, Steffenson BJ (1993) A molecular, isozyme and morphological map of the barley (Hordeum vulgare) genome. Theor Appl Genet 86:705–712Google Scholar
  9. Mackill DJ, Bonman JM (1992) Inheritance of blast resistance in near-isogenic lines of rice. Phytopathology 82:746–749Google Scholar
  10. Paterson AH, Tanksley SD, Sorrells ME (1991) DNA markers in plant improvement. Adv Agron 46:39–89Google Scholar
  11. Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-termination inhibitors. Proc Natl Acad Sci USA 74:5463–5467Google Scholar
  12. Simmons CR, Litts JC, Huang N, Rodriguez RL (1992) Structure of a rice β-glucanase gene regulated by ethylene, cytokinin, wounding, salicylic acid and fungal elicitors. Plant Mol Biol 18:33–45Google Scholar
  13. Tanksley SD, Causse T, Fulton T, Ahn N, Wang Z, Wu K, Xiao J, Yu Z, Second G, McCouch SR (1992) A high density molecular map of rice genome. Rice Genet Newsl 9:111–115Google Scholar
  14. Wang GL, Mackill DJ, Bonman M, McCouch SR, Champoux M, Nelson R (1994) RFLP mapping of genes conferring complete and partial resistance to blast in a durably resistant rice cultivar. Genetics 136:1421–1431Google Scholar
  15. Williams MNV, Pande N, Nair S, Mohan M, Bennett J (1991) Restriction fragment length polymorphism analysis of polymerase chain reaction products amplified from mapped loci of rice (Oryza sativa L.) genomic DNA. Theor Appl Genet 82:489–498Google Scholar
  16. Yu ZH, Mackill DJ, Bonman JM, Tanksley SD (1991) Tagging genes for blast resistance in rice via linkage to RFLP markers. Theor Appl Genet 81:471–476Google Scholar
  17. Zhu L, Chen Y, Ling Z, Xu Y, Xu J (1993) Identification of molecular markers linked to a blast resistance gene in rice. In: You CB, Chen ZL (ed) Agric Biotechnol. China Science and Technology Press, Beijing, China, p 213Google Scholar

Copyright information

© Springer-Verlag 1995

Authors and Affiliations

  • S. Hittalmani
    • 1
  • M. R. Foolad
    • 3
  • T. Mew
    • 1
  • R. L. Rodriguez
    • 2
  • N. Huang
    • 1
  1. 1.International Rice Research InstituteManilaPhilippines
  2. 2.Section of Molecular and Cellular BiologyUniversity of CaliforniaDavisUSA
  3. 3.Department of HorticultureThe Pennsylvania State UniversityUniversity ParkUSA

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