Analysis of Phytophthora parasitica by retrotransposon-derived DNA Fingerprinting

以反轉位子局部序列爲探針進行疫病菌DNA指紋分析

劉瑞芬(Ruey-Fen Liou) ； 李政暾(Jent-Turn Lee) ； 李曉菁(Hsiao-Ching Lee) ； 安寶貞(Pao-Jen Ann)

《Botanical Bulletin of Academia Sinica》 43卷1期 (2002/01) Pp. 21-29

https://doi.org/10.7016/BBAS.200201.0021

摘要

本研究以Tyl-copia反轉位子局部序列爲探針進行疫菌DNA指紋分析。疫病菌可危害之寄主植物種類繁多，重要作物包括柑橘、百合、蘭花及多種觀賞植物都是其危害對象，是台灣重要植物原真菌。研究顯示自菸草、萬看青及枇杷所分離到之疫病菌菌株，不論在型態或病原性都與一般典型疫病菌有顯蓍差異，爲了探討這種差異與疫病菌遺傳變異之關連性，本研究自疫病菌選殖Tyl-copia反轉位子之反轉錄酶基因的局部序列，以之製備探針(G2Ty-1)進行南方雜合分析的研究結果發現，G2Ty-1爲一中度重自複性序列，其雜合圖譜跨五個無性世代維持一致，因此有相當潛力可應用爲分子標記。以G2Ty-1爲探針進行疫病菌DNA指紋分析之結果顯示，雖然不同菌株之G2Ty-1雜合圖譜呈現高度多型性，但不論採集地點是哪裏，自菸草，萬年青及枇杷所分離到之“非典型”疫病菌菌株各顯現出寄主專一性圖譜。進一步根據DNA指紋進行UPGMA群叢分析後發現，可將供試菌株分成三群，其中菸草與萬年青分離株各自形成一個群叢，與其他供試菌株間有明顯的區隔，顯示這些菌株的病原性特化現象有其遺傳基礎，值得進一步深入探討。

關鍵字

反轉位子 ； 疫病菌 ； 基因型 ； DNA指紋分析 ； 寄主專一性

並列摘要

Phytophthora parasitica, being able to attack a wide variety of plants, is a very important plant pathogen in Taiwan. It has been shown that isolates of P parasitica from tobacco, dieffenbachia, and loquat differed significantly from other isolates in morphology and pathogenicity and were recognized as ”atypical” types of the fungus. In the current study, a partial sequence of the reverse transcriptase gene of a Tyl-copia retrotransposon was cloned from P. parasitica and used as the probe, designated herein as G2Ty-1, for genomic Southern hybridization analyses of P. parasitica. It was demonstrated that G2Ty-1 existed as a moderately repetitive sequence in the genome of P. parasitica. The banding pattern of G2Ty-1 was mitotically stable for at least five consecutive asexual generations. DNA fingerprint analysis of P. parasitica using G2Ty-1 as the probe demonstrated that the banding patterns of G2Ty-1 were highly polymorphic among the isolates analyzed. “Atypical” strains from tobacco, dieffenbachia, and loquat, nonetheless, displayed host-specific banding patterns. Phylogram generated by cluster analysis demonstrated that isolates from tobacco and dieffenbachia were well separated from each other and from all other isolates of P. parasitica. These results provided a genetic basis for distinguishing these isolates from others and validate the use of retrotransposon as a genetic marker to study phylogeny in Phytophthora parasitica.

並列關鍵字

DNA fingerprinting ； Genotypes ； Host specificity ； Phytophthora parasitica ； Retrotransposon