水稻抗徒長病基因座之分子定位與育種

Molecular mapping and breeding for loci conferring bakanae resistance in rice

陳思聿(Szu-Yu Chen)

指導教授 ： 鍾嘉綾

國立臺灣大學/生物資源暨農學院/植物病理與微生物學研究所/博士(2020年)

https://doi.org/10.6342/NTU202004325

摘要

水稻為重要糧食作物，由Fusarium fujikuroi引起之水稻徒長病能造成稻米在產量與品質方面的損失，受徒長病菌感染的水稻植株於秧苗期產生複雜病徵，包括徒長、纖細瘦弱、葉片夾角大，嚴重者可導致植株死亡，藉由有效的稻種消毒，徒長病以往得以被良好控制，然而，近幾年於許多亞洲國家皆紛紛報導徒長病之疫情嚴重度增加及抗藥性菌株的出現。為防治水稻徒長病，種植抗病品種為一有效、經濟且環保的可能替代策略。本研究首先自全球核心種原rice diversity panel 1與雙親本雜交族群 (台稉16號x Budda) 中，藉由全基因體關聯定位與連鎖定位尋找徒長病之抗病基因座，共定位出16個抗病基因座，其中qBK1.7區域與已報導之qBK1和qFfR1重疊，並成功以IR64 x Nipponbare族群，透過連鎖定位方式完成驗證，針對20個抗感表現不同的水稻品種於該區域進行序列比對，結果顯示Os01g0601625與Os01g0601675有較高可能性與徒長病抗性相關；利用台稉16號x Budda之166個F9重組自交系進行連鎖定位分析，定位出一個新穎之基因座qBK2.1，及一個與先前報導之qBK1.3位置重疊之qBK1.8。接著，自定位結果與先前之轉錄體分析中挑選出九個候選基因，以即時定量聚合酶連鎖反應或水稻突變體驗證其基因功能性，基因表現量定量結果顯示茉莉酸之訊息傳導可能參與水稻對徒長病菌侵染之反應，且施用甲基茉莉酸能延遲水稻感病品種的病徵發展。徒長病之抗病育種方面，自台農71號 x Budda、台稉16號x IR78581-12-3-2-2、台農71號 x IR78581-12-3-2-2族群中，透過人工接種方式，於F4至F7世代進行抗性選拔，篩選出17個對四株本土代表性徒長病菌菌株具有抗性之水稻品系；而藉由新開發之分子標誌檢測22個臺灣商業品種可能帶有之徒長病抗性，大多數品種於qBK1.4、qBK1.7、qBK2.1、qBK10.1不帶有抗病基因型，這些基因座可作為後續育種改良之標的；為增加田間應用性，亦嘗試建構抗病基因堆疊之品系，逐步取得能同時對徒長病、稻熱病及白葉枯病具抗性之水稻品系。期盼此研究能提供水稻對徒長病抗性研究之基礎，並能為未來防治策略擬定與抗病育種提供助益。

關鍵字

水稻徒長病 ； Fusarium fujikuroi ； 全基因體關聯定位 ； 連鎖定位 ； 抗病育種

並列摘要

Rice is an important staple food crop. Bakanae disease caused by Fusarium fujikuroi caused considerable reduction in rice yield and quality. Infected rice plants showed complex symptoms such as overgrowth, slender, a large leaf angle, and even death at seedling stage. Seed disinfection with chemical fungicides was effective for bakanae control for a long time. However, in recent years, the increasing epidemics of bakanae disease and emergence of fungicide-resistant fungal isolates were reported in several Asian countries. Use of resistant cultivars can be an effective, economic, and eco-friendly alternative control approach for bakanae disease control. In the first part of this study, loci conferring resistance to bakanae disease were identified using rice diversity panel 1 and Taikeng16 x Budda population by genome-wide association mapping and linkage mapping, respectively. A total of 16 quantitative trait loci (QTLs) were mapped. A candidate QTL (qBK1.7) was co-localized with the previously reported qBK1 and qFfR1 and validated by linkage analysis of a recombinant inbred line (RIL) population derived from IR64 x Nipponbare. Full-length sequencing and comparison of qBK1.7 in 20 rice accessions indicated that Os01g0601625 and Os01g0601675 may control bakanae resistance. Linkage mapping conducted using a population of 166 F9 RILs derived from Taikeng16 x Budda identified two resistant QTLs: qBK2.1 is a novel QTL and qBK1.8 was overlapped with the previous reported qBK1.3. Secondly, nine candidate genes selected based on QTL mapping and previous transcriptome analysis were verified using real-time quantitative reverse transcription PCR (qRT-PCR) or transgenic rice lines. The qRT-PCR result showed that jasmonic acid signaling may involve in rice in response to F. fujikuroi infection and the application of methyl jasmonate delayed bakanae disease development in susceptible cultivar (Zerawchanica karatals). Finally, resistance breeding for bakanae disease was conducted. F4 to F7 progeny lines from the crosses of Tainung 71 (TNG71) x Budda, TK16 x IR78581-12-3-2-2, and TNG71 x IR78581-12-3-2-2 were subjected to artificial inoculation and resistance selection. Total 17 lines resistant to four representative local isolates of F. fujikuroi were obtained. For future resistance breeding, potential bakanae resistance in 22 Taiwan cultivars was examined using newly developed molecular markers. Most cultivars did not carry resistance alleles at qBK1.4, qBK1.7, qBK2.1, and qBK10.1. These QTLs are proper target for resistance improvement. Moreover, breeding for pyramiding lines for resistance to bakanae disease, rice blast, and bacterial blight is underway. We hope that this study can provide the fundamental basis of bakanae resistance to facilitate the development of control strategy and future resistance breeding.

並列關鍵字

bakanae disease ； Fusarium fujikuroi ； genome-wide association mapping ； linkage mapping ； resistance breeding

參考文獻

Abdurakhmonov IY, Abdukarimov A (2008) Application of association mapping to understanding the genetic diversity of plant germplasm resources. International journal of plant genomics 2008:1–18

Agrawal GK, Rakwal R, Jwa N-S (2000) Rice (Oryza sativa L.) OsPR1b gene is phytohormonally regulated in close interaction with light signals. Biochemical and Biophysical Research Communications 278:290-298.

Agrawal GK, Rakwal R, Jwa N-S, Agrawal VP (2001) Signalling molecules and blast pathogen attack activates rice OsPR1a and OsPR1b genes: A model illustrating components participating during defence/stress response. Plant Physiology and Biochemistry 39:1095-1103.

Ali ML, McClung AM, Jia MH, et al (2011) A rice diversity panel evaluated for genetic and agro-morphological diversity between subpopulations and its geographic distribution. Crop Science 51:2021-2035.

Amatulli MT, Spadaro D, Gullino ML, Garibaldi A (2010) Molecular identification of Fusarium spp. associated with bakanae disease of rice in italy and assessment of their pathogenicity. Plant Pathology 59:839-844.