水稻臺中秈10號及臺稉2號抗稻熱病基因座之鑑定

Abstract

稻熱病由Pyricularia oryzae (syn. Magnaporthe oryzae) 所引起，為限制水稻生產的重要病害之一。為降低已知抗病基因於田間被病原菌突破的風險，尋找新抗病基因、育成新品種及合理佈署抗病基因組合，為水稻抗病育種的重要策略。臺中秈10號為臺灣秈稻栽培面積最大的品種，於臺灣田間長期保持對稻熱病的優良抗性。臺稉2號為臺灣主流之栽培品種，但已喪失田間抗性。為釐清兩品種抗稻熱病之遺傳基礎，本研究首先以22株採集自臺灣不同地區及年份的P. oryzae菌株接種臺中秈10號與臺稉2號。臺中秈10號對所有測試菌株皆呈現抗病反應，而臺稉2號則對其中16株表現抗病性。進一步挑選於兩親本間表現親和性差異之菌株20TD-IRRI14-1-1與HL4a1-2010，接種「臺稉2號 × 臺中秈10號」所衍生之245個重組自交系 (recombinant inbred lines, RILs)，以定位抗稻熱病相關之數量性狀基因座 (quantitative trait loci, QTLs)。本研究利用double digest restriction-associated DNA sequencing (ddRAD-seq) 獲得之2,761個single nucleotide polymorphisms (SNPs) 分子標誌，建構全長為1,453.7 cM之遺傳圖譜，並以連鎖分析定位出一個主效QTL及一個微效QTL。由兩菌株接種結果偵測到的主效QTL位於第12號染色體7.2–13.1 Mb與9.4–10.8 Mb重疊區間內，其logarithm of odds (LOD) 值分別為17.5與19.7，可降低罹病級數2級與3級。微效QTL僅於HL4a1-2010接種結果中被偵測到，位於第5號染色體0.1–2.9 Mb區間 (LOD值為3.9)，可降低罹病級數1級。經由Sanger定序分析兩親本於已知抗稻熱病基因之序列，臺中秈10號攜帶抗性基因Ptrb與Pik-s，而Pi2/9則鑑定出功能尚待釐清之新穎等位基因序列；臺稉2號則攜帶Piz-t與Pik-s。針對50個臺灣水稻栽培品種之Ptr單倍型進行解序並結合嘉義及臺東病圃資料，發現在2008年至2025年間於病圃中表現穩定抗性的品種皆攜帶抗病Ptr單倍型 (PtrA及Ptrb)。PtrA主要存在於稉稻品種，Ptrb多分布於秈稻品種，未檢測到任何栽培種攜帶另一抗病單倍型PtrB。綜合上述結果，推測臺中秈10號之抗性主要源於Ptrb等位基因，該位點 (10.8 Mb) 與QTL定位中偵測到的主效QTL區段重疊。臺中秈10號亦攜帶Pik-s及第5號染色體上的微效QTL，有助於其在田間表現持久抗性。本研究成果可作為未來水稻品種輪替及抗稻熱病育種策略擬定之重要依據。

Rice blast, caused by Pyricularia oryzae (syn. Magnaporthe oryzae), is one of the most destructive diseases limiting rice production worldwide. To mitigate the risk of resistance breakdown of deployed resistance genes in the field, the identification of novel resistance genes, development of new cultivars, and rational deployment of resistance gene combinations are essential strategies in rice breeding programs. Taichung Sen No. 10 (TCS10) is the most widely cultivated indica rice variety in Taiwan, and it has maintained durable blast resistance under field conditions for decades. Taikeng No. 2 (TK2), a major japonica cultivar, has lost its field resistance. To elucidate the genetic basis of blast resistance in these two cultivars, 22 P. oryzae isolates collected from different regions and years in Taiwan were inoculated on TCS10 and TK2. TCS10 exhibited resistant reactions to all tested isolates, whereas TK2 was resistant to 16 of them. Two isolates, 20TD-IRRI14-1-1 and HL4a1-2010, showing contrasting compatibility between the parental cultivars, were selected to inoculate a population of 245 recombinant inbred lines (RILs) derived from TK2 × TCS10 for quantitative trait loci (QTL) mapping. Using 2,761 single nucleotide polymorphism (SNP) markers obtained from double digest restriction-associated DNA sequencing (ddRAD-seq), a genetic linkage map spanning 1,453.7 cM was constructed. QTL analysis identified one major-effect QTL and one minor-effect QTL associated with blast resistance. The major-effect QTL detected from the two isolates were located on chromosome 12 within the overlapped intervals of 7.2–13.1 Mb and 9.4–10.8 Mb, with logarithm of odds (LOD) scores of 17.5 and 19.7, respectively, reducing the disease severity index by 2 and 3 scales. A minor-effect QTL was detected only from the HL4a1-2010 inoculation, located on chromosome 5 (0.1–2.9 Mb; LOD = 3.9), contributing to a 1-scale reduction in disease severity index. Sanger sequencing of known blast resistance genes revealed that TCS10 carries the resistance genes Ptrb and Pik-s, while its Pi2/9 locus harbors a novel allele of unknown function; TK2 carries Piz-t and Pik-s. Haplotype analysis of the Ptr locus in 50 Taiwanese rice cultivars, combined with Chiayi and Taitung blast nursery data from 2008 to 2025, demonstrated that cultivars exhibiting stable field resistance consistently carry resistant Ptr haplotypes (PtrA or Ptrb). PtrA was predominantly found in japonica cultivars, whereas Ptrb was mainly distributed in indica cultivars; no cultivar carrying the resistant PtrB haplotype was identified. Collectively, these results suggest that the resistance of TCS10 is primarily attributable to the Ptrb allele, which is located at 10.8 Mb on chromosome 12 and overlaps with the major-effect QTL identified in this study. TCS10 also carries Pik-s and a minor-effect QTL on chromosome 5, which likely contribute to its long-term field resistance. These findings provide an important basis for future rice cultivar rotation and the development of blast resistance breeding strategies.