- 學位論文
香蕉萎縮病病毒增額複製酶基因體生物學及分子生物學特性分析
Biological and molecular characterization of additional replicase encoded components associated with Banana bunchy top virus
摘要
香蕉為世界上重要糧食作物之一，亦是台灣地區的重要果樹。而香蕉萎縮病是香蕉重要的病毒病害，影響產業甚鉅。香蕉萎縮病的病原為香蕉萎縮病病毒 (Banana bunchy top virus, BBTV) ，屬於Nanoviridae科，Babuvirus屬之球狀病毒。BBTV的基因體主要由六個環形的單鏈DNA所組成，稱為integral components。但在某些系統中除integral components外，尚有其他會自行複製之基因體存在，稱為additional Reps，目前發現了八個環形單鏈DNA，分別為Y、W1、W2、W3、W4、S1、S2和S3。為探討additional Reps在BBTV感染過程中所扮演的角色，本實驗收集香蕉萎縮病之罹病株，並利用additional Reps之專一性引子對進行偵測，並將這些病株依照體內之additional Reps存在的情況共分為五群，每群中選一代表株，再以蕉蚜 (Pentalonia nigronervosa) 將這六棵病株之病毒傳至健康植株上，觀察其病徵，初步發現若BBTV感染株內未含有任何additional Reps時其病徵最為輕微，若感染株內含additional Reps時其病徵均較為嚴重。其中以含有S1或Y與S2時，香蕉萎縮病的病徵最為嚴重，而感染僅有Y存在時，病徵相對來說較為輕微。另外，藉由親緣演化分析Nanoviridae之additional Reps，則發現BBTV additional Reps由一共同祖先演化而來，並朝兩個方向演化。同時，在open reading frame (ORF) 區域內的stem-loop right site common region (SRCR) 是於蛋白質層次的一重要motif。為進一步釐清BBTV integral components與additional Reps之間的交互作用，並真正明瞭這些additional Reps的功能與其存在對BBTV的影響，因此構築integral components DNA 1-6和additional Reps Y與S1的1.2至1.6倍全長之感染性克隆 (infectious clones) 。並在香蕉懸浮細胞內，證實DNA 1的infectious clone能夠複製，為BBTV及additional Reps的研究奠定重要的基礎。
並列摘要
Banana is one of the most important fruit crops around the world. Banana bunchy top disease caused by Banana bunchy top virus (BBTV) is a destructive disease for banana. BBTV is isometric and belongs to the genus Babuvirus, in the family Nanoviridae. BBTV consists of six circular single-stranded integral DNA components, DNA 1-6. In some isolates, however, in additional to the integral components other replication competent components (additional Reps) are also associated with BBTV infection. Currently, 8 different additional Reps, Y, W1, W2, W3, W4, S1, S2, and S3, have been reported. To understand the roles of BBTV additional Reps in BBTV infection, we collected samples from field and using primer pairs specific to each BBTV additional Reps to detect the additional Reps. The samples can be grouped into five groups by the presence of additional Reps in a single infected plant. One isolate were selected from each group and use as source to inoculate healthy bananas by using of banana aphids (Pentalonia nigronervosa). Briefly, BBTV associated with S1, or Y plus S2 induced the most severe symptoms, and BBTV associated with Y induced comparatively slight severe symptoms. In addition, BBTV with no additional Reps association induced less severe symptoms. Phylogenetic analysis of additional Reps ORFs (open reading frames) encoded by Nanoviridae reveled that BBTV additional Reps evolved from a common ancestor and later evolved to two directions and form 6 monoclade. And we find that SRCR (stem-loop right site common region; a specific region conserved among BBTV additional Reps) within ORF evolved with a strong selection pressure; which suggest this region contains an unidentified protein motif. To further study the interaction between integral components and additional Reps, we construct BBTV integral components DNA 1-6 and additional Reps Y and S1. Each infectious clone consists of greater-than-unit (range 1.2 to 1.6 mers). We have demonstrated the replication of BBTV DNA 1 infectious clones in banana suspension cells.
參考文獻
延伸閱讀
- 邱建翔（2014）。香蕉ACC氧化酶基因默化轉殖株中果實後熟相關基因表現之分析〔碩士論文，國立臺灣大學〕。華藝線上圖書館。https://doi.org/10.6342/NTU.2014.10389
- 黃怡菁、許圳塗、馬溯軒（1999）。香蕉AAA基因組華蕉系在亞群及AAB品種胚性細胞懸浮培養之生長週期及循環生長特性。中國園藝，45(2)，130-143。https://doi.org/10.6964/JCSHS.199906.0130
- 廖玉琬、徐善德、黃鵬林（1999）。香蕉轉殖細胞利用葡萄糖苷酸酶活性組識化學染色之改進。中國園藝，45(3)，239-244。https://doi.org/10.6964/JCSHS.199909.0239
- 張馨元（2011）。香蕉萎縮病毒及香蕉條紋病毒甲基化之分析與比較〔碩士論文，國立臺灣大學〕。華藝線上圖書館。https://doi.org/10.6342/NTU.2011.01774
- 胡慧琳（2012）。Gene Expression and Promoter Activity Analysis of MhCTR1 Related to Ethylene Signal Transduction in Banana〔博士論文，國立臺灣大學〕。華藝線上圖書館。https://doi.org/10.6342/NTU.2012.11017