Polymorphism analysis of genomic regions associated with broad-spectrum effective blast resistance genes for marker development in rice
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Cultivated European rice germplasm is generally characterized by moderate to high sensitivity to blast, and blast resistance is therefore one of the most important traits to improve in rice breeding. We collected a panel of 25 rice genotypes containing 13 broad range rice resistance genes that are commonly used in breeding programs around the world: Pi1, Pi2, Pi5, Pi7, Pi9, Pi33, Pib, Pik, Pik-p, Pita, Pita 2 , Piz and Piz-t. The efficiency of the selected Pi genes towards Italian blast pathotypes was tested via artificial inoculation and under natural field infection conditions. To characterize haplotypes present in the chromosomal regions of the blast resistance genes, a polymorphism search was conducted in the sequence regions adjacent to the blast resistance by examining DNA from the Pi gene donors with a panel of 5–7 potential receivers (cultivated European rice genotypes). Seven InDel and 8 presence/absence polymorphisms were directly detected by gel analysis after DNA amplification, while sequencing of 12.870 bp through 32 loci in different genotypes revealed 85 SNP (one SNP every 151 bp). Seven SSRs were additionally tested revealing 5 polymorphic markers between donors and receivers. Polymorphisms were used to develop 35 PCR-based molecular markers suitable for introgressing of Pi genes into a set of the European rice germplasm. For this last purpose, allelic molecular marker variation was evaluated within a representative collection of about 95 rice genotypes. Polymorphic combinations allowing introgression of the broad spectrum resistance genes into a susceptible genetic background have been identified, thus confirming the potential of the identified markers for molecular-assisted breeding.
KeywordsRice (Oryza sativa) Blast (Magnaporthe oryzae) resistance Haplotype analysis Marker-assisted selection (MAS) Gene pyramiding
The authors wish to thank the following persons for the gift of rice genotypes and blast isolates: Dr. Harold Bockelman (USDA, Agricultural Research Service, National Small Grains Research Facility, National Small Grains Collection, USA) for C101LAC, Bala, Carreon, Te Qing, Saber, Katy, Kaybonnet, Zenith, Jefferson; Dr. Didier Tharreau (UMR BGPI, CIRAD, France) for C101LAC, IR64, IR1529, K1, K60 and for the three Italian blast strains; Dr. Jong-Seong Jeon (Graduate School of Biotechnology & Plant Metabolism Research Center Kyung Hee University Korea) for RIL260, IRBL5-M, IRBL7-M; Dr. Kazutoshi Okuno (National Institute of Agrobiological Sciences NIAS, Japan) for Kanto 51, Kusabue, BL1, K1, Fukunishiki, Toride 1 and Dr. Guo-Liang Wang (Department of Plant Pathology, The Ohio State University, USA) for 75-1-127 and C101A-51. This study acknowledges financial support of the project VALORYZA (Italian Ministero per le Politiche Agricole e Agroalimentari, DM 301/7303/06), CEREALAB and SITEIA (Italian Region Emilia Romagna) and of the EU co-funded project EURIGEN (049 AGRI GEN RES).
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