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標題: 伏寄普抗性生物型牛筋草之分子抗性機制研究
Study on the molecular resistance mechanism of the fluazifop-resistant biotype of goosegrass [Eleusine indica (L.) Gaertn.]
作者: 林鈺荏
Lin, Yu-Ren
關鍵字: 牛筋草
molecular mechanism
出版社: 農藝學系所
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摘要: 牛筋草[Eleusine indica (L.) Gaertn.]為臺灣蔬菜田、果園等旱地,在高溫季節所發生極為普遍之一年生禾本科雜草。伏寄普(fluazifop-P-butyl)為抑制乙醯輔酶A羧化酵素(acetyl-CoA carboxylase, ACCase)之選擇性萌後除草劑,可有效防除禾本科雜草。近幾年,在臺灣中、南部田區發生牛筋草抗伏寄普現象,造成防治困難。本研究以對伏寄普具抗、感性之牛筋草生物型為材料,比較其形態、生理反應以及選殖、定序比較兩者在DNA序列及羧基轉移酵素功能域(carboxyl transferase domain, CT domain)之胺基酸序列差異,探討其抗性之分子機制。抗、感性牛筋草二者的外表型自幼苗至成熟植株皆極為相似,無法分辨出差異,但由SSR分子標誌多型性分析得知,抗、感生物型基因體之間存在著11.0%的差異性。抗性生物型在伏寄普處理後7天,會恢復生長並顯著加速分蘖,由外表形態傷害指數的劑量反應(dose response)分析得知,抗性生物型(ED50=0.197 mM)與感性生物型(ED50=0.004 mM)之間的抗、感比達49倍,且抗性生物型抗性能力穩定,不受生育階段影響。另外抗性生物型葉綠素含量(1.2 mg/g FW)顯著高於感性生物型(0.1 mg/g FW);而丙二醛(malondialdehyde, MDA)含量(24.9 nmole g-1 FW)則比感性生物型低(35.2 mg/g FW),顯示抗性生物型的細胞受到伏寄普傷害較低。基因選殖和序列比對結果顯示,CT domain之DNA序列發生點突變由腺嘌呤(adenine)轉變為鳥糞嘌呤(guanine),造成位在相對於大穗看麥娘(Alopecurus myosuroides Huds.) CT domain胺基酸序列(AJ310767)的2,078位置上胺基酸,由天門冬胺酸(aspartic acid, D)變成了甘氨酸(glycine, G)。推測此一胺基酸的置換,造成蛋白三級構型大幅度的改變而產生缺口,使得除草劑無法順利與CT domain結合,為牛筋草產生抗性現象的主因。
Goosegrass, a common annual poaseas weed of vegetible fields, orchards and none-cultivated lands, they grows fast in summer time of Taiwan. Fluazifop-P-butyl, a selective post-emergence herbicide, controls the grass effectively by inhibiting the acetyl-CoA carboxylase (ACCase) which is the crucial enzyme of fatty acid biosynthesis. In recent years, many cases of goosegrass were reported to be resistant to fluazifop, and become a big problem in its control especially in the south central fields of Taiwan. In this study, fluazifop resistant and susceptible biotypes of goosegrass are AOPPlied to study the molecular mechanism of resistance by comparing the agronomic traits, pysiological reactions and sequences of ACCase gene. Though the agronimic traits are almost identical between two biotypes, their genomes show 11.0% differences by the analysis of SSR markers. The resistant biotype could significantly regrowth with new tillers at 7 days after fluazifop treatment and the responses (ED50) of resistant and susceptible biotype seedling are 0.197 mM and 0.004 mM, respectively, and the resistance index [ED50 (R) / ED50 (S)] is about 49. The resistance index maintained through the whole growth stages susgests that the resistance is very stable. Besides, comparison of the chorophyll and malondialdehyde (MDA) contents of the resistant biotype (1.2 mg/g FW and 24.9 nmole/g FW) and the susceptible biotype (24.9 nmole/g FW and 35.2 mg/g FW) after treated with 0.5 mM fluazifop, showed that the R biotype is less damaged by fluazifop. Sequences aomparison of the CT domain s found a point mutation (A→G) leading to an amino acid changed (Asp→Gly) at the 2,078 position which may alter the three-dimensional conformation of fluazifop active site, and cause the resistance.
其他識別: U0005-0808201320211000
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