- 學位論文
黃楊花部器官決定同源基因的鑑定與演化
Identification and evolution of floral organ identity gene homologues in Buxus microphylla ssp. sinica (Buxaceae)
摘要
目前經由模式植物阿拉伯芥的研究，建立了花部器官決定基因的調控（ABCDE 模組），即花萼由A、E 群基因共同決定，花瓣由A、B、E 群基因，雄蕊由B、C、E 群基因，而心皮由C 和E 群基因決定。A、B、C、E 四群基因在被子植物的演化過程中發生過數次的複製事件，其中比較重要的一次發生在早期被子植物演化出來之時，形成B/C/D/E 群基因的次系群；另一次複製事件則發生在真雙子葉植物基群（Basal eudicots）附近，形成僅見於核心真雙子葉植物（Core eudicots）的A/B/C/D/E 群基因之次系群。根據前人的譜系關係研究，黃楊科很接近真雙子葉植物基群第二次主要複製事件的時間點，因此鑑定黃楊科植物的花部基因，將有助於釐清調控花部器官形成基因的演化情形。黃楊（Buxus microphylla ssp. sinica）是本篇研究所使用的植物材料，共有9 個花部器官同源基因在本研究中成功被鑑定出來。其中有2 個屬於A 群基因，4 個屬於B 群基因，2 個C 群基因以及1 個E 群基因。從RT-PCR 的結果可以看到這些花部基因廣泛地表現在花部各輪器官，並不表現於葉部。唯一的例外是BumPI2，其在假雌蕊缺乏表現，而在雄蕊僅微弱表現，與其他基因的表現情形較不相同。黃楊花部器官決定基因的表現和現行的ABCDE 模組並不一致，顯示黃楊與模式植物的調控模式可能不同，但尚待進一步的研究以進行確認。另外，根據譜系分析的結果，鑑定出的黃楊花部基因分別隸屬於ABCE 不同的基因系群內，位於真雙子葉植物之中。黃楊的花部基因總是和三角咪屬或者昆欄樹屬的基因分成一群，在靠近核心真雙子葉植物的位置形成姊妹群。BumAP3-1/2/3 是分析結果中的例外，有時會和三角咪屬（Pachysandra）植物的基因一起在核心真雙子葉植物中成群，成為euAP3 基因類群的姊妹群。由序列的分析亦顯示，框移突變的模擬可以推斷黃楊的基因有高度框移突變的潛力，而框移突變可能是在基因複製事件之後導致核心真雙子葉植物分化的重要機制。
並列摘要
Floral organ determination is best explained by the ABCDE model postulated by genetic studies of Arabidopsis thaliana. Sepals are determined by A and E class genes; petals are determined by A, B, and E; stamens by B, C, and E; and carpels by C and E class genes. A, B, C, and E class gene lineages are known having duplicated several times during the evolution of angiosperms. One of the noted major duplication events occurred in the origin of the early angiosperms, leading to the formation of subgroups of B/C/D/E class. Another one occurred near the basal eudicots and gave rise to further subgroups in A/B/C/D/E class genes among core eudicots. The phylogenetic position of the family Buxaceae is located right where the second major duplication of ABCDE genes might have occurred, which is supported by multiple gene (nuclear 18S rDNA, chloroplast rbcL and atpB) phylogenetic analyses. Therefore, the identification and characterization of floral organ identity genes in Buxaceae is critical in elucidating the gene evolution related to floral organ formation. Floral organ identity gene homologues of Buxus microphylla ssp. sinica were screened, and nine gene homologues of B. microphylla ssp. sinica were successfully identified in this study. Two of them are A class gene homologues, four B class, two C class, and one E class. Expression patterns based on RT-PCR of floral organ identity genes of Buxus do not match with current ABCDE model. There is no expression of all identified genes in leaves, and all genes are expressed in all floral parts except BumPI2. BumPI2 does not express in pistillodes, and only weakly expresses in stamens. Whether or not the discrepancy is due to a deviation from the standard floral model awaits further examinations. According to the phylogenetic analyses, the homologous genes of Buxus belong to their expected clades, showing a position near the base of core eudicots. Genes of Buxus are always sister to genes of Pachysandra or Trochodendron,and this clade is close to core eudicots in all analysis. BumAP3 genes are the only exception that sometimes grouped with Pachysandra genes as sister group to euAP3 clade. Moreover, frameshift modulation of BumAP3-1/2/3 genes indicates their high frameshift potential and frameshift mutation might be the key mechanism occurred after duplication event and led to diversification of core eudicots.