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題 名 | 以半巢式PCR偵測樹枝與預採果實中的檬果蒂腐菌=Detection of Quiescent Infection of Mango Stem End Rot Pathogen Lasiodiplodia Theobromae in Shoot and Pre-plucked Mango Fruit by Seminested PCR |
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作 者 | 林玉儒; 王嬿婷; 楊宏仁; 汪碧涵; | 書刊名 | 植物病理學會刊 |
卷 期 | 18:4 2009.12[民98.12] |
頁 次 | 頁225-235 |
分類號 | 435.325 |
關鍵詞 | 檬果蒂腐病; 黑色蒂腐菌; 半巢式PCR; 監測技術; Stem end rot of mango; Lasiodiplodia theobromae; Semi-nested PCR; Monitoring technique; |
語 文 | 中文(Chinese) |
中文摘要 | 本研究由樣果罹病果實上分得55株菌株,經鑑定為Botr)叫phaeriadothidea, B. ribis, Pestalotiopsis 亭,Phomopsis 亭,Colletotrichum acutatum, C.gloeosporioid,凹,其中6株以形態與 核酸序列鑑定為樣果蒂腐菌Lasiodiplodiatheobromae Griffon & Maubl.。由L. theobromae核醋 體DNA的ITS區域設計種專一性可|子Lth1'與廣效性引子ITS4配對,以最適煉合溫度580C增幅得到一約420bp的產物,上述其他菌種不會形成產物。為提升偵測靈敏度,建立半巢式 PCR:第一段以ITS5/ITS4廣效性引子對增幅,第二段續以Lthl/ITS4增幅,可使偵測靈敏度達 125 fg L.theobromae全量DNA 02004至2005年間,將半巢式PCR 1貞測技術應用於田間愛文 檬果枝條帶菌調查與預踩果實檢測。結果發現,三月至五月,病原菌由舊枝侵入當年度新枝,進入果慣,舊校中棲息的蒂腐菌為重要的果實採收後蒂腐病的感染源。當年度新技基部偵測率可達45.59忘,新舊枝交接處(63.39毛)與老枝頂端(72.7仰的偵測率最高,老枝15公分處仍能測得蒂腐菌。由屏東、台南和嘉義等樣果主要產區的七個果園,於採收前1-1.5個月收集預採果實,萃取果蒂及其周圍果皮組織DNA'再將預採果實浸潤生長素益收(ethylene)以催熟,調查罹病率。2004年,樣果果實蒂腐病罹病率在09毛�829忘,半巢式PCR偵測率則在6.39毛�769毛之間;2005年果實罹病率約為20%�92.99忘,半巢式PCR偵測率則約29.29毛�81% 0雖然各果園半 巢式PCR偵測率與預採果實罹病率不盡相符,但罹病率超過109毛的樣本,半巢式PCR偵測率皆超過109忘。依據政府��T藏期果實病害預測方法r訂定凡經預測之果園,蒂腐病罹病果實率在109毛以下,得選為日本外銷果園。半巢式PCR偵測以其靈敏與快速的優點,適宜開發成監測與偵測的技術。 |
英文摘要 | A seminested PCR-based method was developed for specific detection of mango (Mangifera indica L.) stem end rot pathogen Lasiodiplodia theobromae Griffon & Maubl. (=Botryodiplodia theobromae Pat.) in plant tissues. A specific primer Lth1 was designed from rDNA internal transcribed spacers (ITS) region of L. theobromae. Under stringent PCR conditions, a 420-bp amplicon formed from L. theobromae DNA but not from other stem end rot pathogens such as Botryosphaeria dothidea, B. ribis, Pestalotiopsis sp., Phomopsis sp., Colletotrichum acutatum, and C. gloeosporioides. Seminested PCR using primer pairs of ITS5/ ITS4 and Lth1/ITS4 was sensitive enough to detect 125 fg of genomic DNA. The technique was used to detect L. theobromae and study the colonization of the pathogen in shoots and pre-plucked fruits. The pathogen in asymptomatic old branches was detected by seminested PCR. It survived endophytically. During March to May, it grew into new branch and extended to inflorescence. The endophytic pathogen seems to be an important inoculum source of the post-harvest disease. In infected orchards, the pathogen was detected in 45.5% of the base of new shoots, in 63.2% tissues between new shoot and old branch, and in 72.7% old branch top. Pre-plucked mango samples were collected from 7 orchards in Pingtung, Tainan, and Chiayi. The pathogen in fruit stalk and the skin near stalk was detected by seminested PCR. The preplucked samples were treated with ethylene and stem end rot symptoms appeared in 9-12 days, significantly after 12 days. In 2004, 0%-82% fruit samples developed stem end rot and the detection rate of L. theobromae was between 6.3%-76%. In 2005, the disease rate of the fruit was 20%-92.9% and the detection rate was about 29.2%-81%. The detection rate of pre-plucked samples was not always consistent with the disease rate. However, when the detection rates in these samples were higher than 10% by seminested PCR, their disease rates were higher than 10%. Based on the standards of mango export orchards, the mango stem end rot disease rates need to be lower than 10%. The seminested PCR assay showed promise as a monitor tool for the mango stem end rot pathogen. |
本系統中英文摘要資訊取自各篇刊載內容。