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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 王俊能 | |
| dc.contributor.author | Meng-Jung Ho | en |
| dc.contributor.author | 何孟容 | zh_TW |
| dc.date.accessioned | 2021-06-16T16:42:05Z | - |
| dc.date.available | 2015-08-27 | |
| dc.date.copyright | 2012-08-27 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-08-27 | |
| dc.identifier.citation | References
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/63443 | - |
| dc.description.abstract | 在苦苣苔科 (Gesneriaceae) 中，子葉不等大為其特有的現象。在種子萌芽後，其中一片子葉基部的分生組織 (basal meristem) 會持續不斷生長成大子葉，另一小子葉則逐漸停止生長、最後凋亡。在前人研究中，施加吉貝素 (GA3) 會抑制basal meristem的活力，使菫蘭不等大子葉的現象消失，形成二片小子葉的型態，這和一般植物中維持頂端分生組織活力需要有低濃度的吉貝素，似乎有著相似的模式。在本研究中，我們藉由吉貝素處理後與正常型堇蘭的形態觀察差異為基礎，進行RT-PCR、RNA原位雜交的試驗，嘗試解開吉貝素在大小子葉中所扮演的角色。我們分離出在菫蘭 (Streptocarpus rexii) 中降解吉貝素的酵素基因SrGA2ox以及合成吉貝素的酵素基因SrGA20ox；在堇蘭幼苗形成不等大子葉的時期，SrGA2ox表現在basal meristem以及groove meristem (形成新葉phyllomorph及長成花序的分生組織) 的位置，證實basal meristem的形成是需要SrGA2ox的表現降解吉貝素，以低濃度的吉貝素來維持basal meristem的活力，而產生持續不斷生長的大子葉；SrGA20ox則表現在小子葉和大子葉的尖端，推測可能參與細胞分裂區外的細胞生長部份。此外，我們也檢測菫蘭的胚胎時期，吉貝素降解基因SrGA2ox表現在二子葉及子葉的近軸面，意味著二片子葉皆具有分生組織的能力，所以大小子葉現象是種子萌芽後才受到環境調控形成；而吉貝素合成基因SrGA20ox在胚胎時期集中表現在shoot meristem (兩片子葉中間) 的位置和子葉的遠軸面，可能因累積了高濃度的吉貝素在將來莖頂分生組織位置，使植株無法形成正常頂芽現象。因為細胞分裂素（CK）處理會使大小子葉現象消失但相反的形成兩片大子葉現象，未來可繼續研究細胞分裂素在堇蘭子葉生長過程中所扮演的角色，試著去解開吉貝素及細胞分裂素與苦苣苔科植物不等大子葉如何發育的謎團。 | zh_TW |
| dc.description.abstract | Anisocotyly is an unorthodox phenomenon among angiosperms, found only in the family members of Gesneriaceae. The rosulate species Streptocarpus rexii exhibits an anisocotylous growth pattern as one of its cotyledons (macrocotyledon) grows continuously via sustained basal meristem (BM) activity at the proximal end. Previous findings suggest that exogenous gibberellin (GA3) can inhibit anisocotyly via suppression of the formation of the BM, causing two equal-sized, small microcotyledons. This study is therefore aimed to shed light on the role of gibberellin (GA) in anisocotyly by revealing the expression pattern of GA catabolism and biosynthesis genes, SrGA2ox and SrGA20ox, to determine how they regulate the unusual basal meristem activity that gives rise to anisocotyly in S. rexii. RT-PCR and RNA in situ hybridization demonstrated that in the macrocotyledon, SrGA2ox shows restricted expression in the basal meristem and also in a shoot apical meristem (SAM) equivalent groove meristem (GM), where further phyllomorph and inflorescence are produced. On the contrary, SrGA20ox transcripts, distributed in the microcotyledon and the distal end of the macrocotyledon, never distributed in the meristem area. These results show agreement with that of model plants, which also suggest that a low concentration of GA is necessary for maintaining meristem activity. Moreover, this mutually exclusive expression pattern between SrGA2ox and SrGA20ox was established since the embryo stage. SrGA2ox was expressed on the adaxial side of both cotyledons, whereas SrGA20ox was distributed in the shoot meristem (SM) and abaxial side of the cotyledons during the embryo stage. The expression of SrGA20ox in the SM may explain the lack of normal shoot initiation in S. rexii seedlings. Because it is also known that exogenous cytokinin (CK) can induce both cotyledons into macrocotyledons, further investigation on the expression profiles of CK genes and their interaction with GA genes is important to fully uncover the developmental mechanism of anisocotyly. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T16:42:05Z (GMT). No. of bitstreams: 1 ntu-101-R99b44012-1.pdf: 2015950 bytes, checksum: 53881a4a70c0cbf5d7917333149fced6 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | Content
Acknowledgment...................................................................................................................i 中文摘要...............................................................................................................................ii Abstract................................................................................................................................iii Content.................................................................................................................................iv Index of Figures and Tables..............................................................................................vii Chapter 1. Introduction.......................................................................................................1 1.1 The uniqueness of Streptocarpus rexii............................................................................1 1.2 Phytohormones’ role in anisocotyly................................................................................3 1.3 Maintaining the meistem activity in model plants...........................................................4 1.4 KNOX gene in Streptocarpus rexii..................................................................................6 Chapter 2. Materials and Methods....................................................................................10 2.1 Plant materials................................................................................................................10 2.2 Plant-hormone treatment................................................................................................10 2.3 Assessment of meristematic activity..............................................................................11 2.3.1 Cell division observation.............................................................................................11 2.3.2 Meristem positions ascertainment.................................................................12 2.4 Cloning of GA2-oxidase and GA20-oxidase genes homologs from Streptocarpus rexii12 2.4.1 Isolation of SrGA2ox gene...........................................................................................13 2.4.1.1 RNA extraction.........................................................................................................13 2.4.1.2 First strand cDNA synthesis.....................................................................................14 2.4.1.3 SrGA2ox isolation.....................................................................................................15 2.4.2 Isolation of SrGA20ox gene.........................................................................................15 2.4.2.1 DNA extraction........................................................................................................15 2.4.2.2 SrGA20ox isolation..................................................................................................16 2.4.3 Standard polymerase chain reaction (PCR).................................................................16 2.4.4 Homologous assessment of the SrGA2ox and SrGA20ox genes.................................17 2.4.4.1 Phylogenetic tree construction of SrGA2ox and SrGA20ox.....................................17 2.4.5 3’ rapid amplification of the DNA ends (3’ RACE)...................................................17 2.5 Reverse transcription-PCR (RT-PCR)...........................................................................18 2.6 In situ hybridization.......................................................................................................18 2.6.1 Fixation and embedding of samples...........................................................................19 2.6.2 Preparation of RNA probes.........................................................................................20 2.6.3 Preparing for in situ hybridization..............................................................................25 2.6.4 Section, deparaffinization and rehydration.................................................................25 2.6.5 Pre-hybridization........................................................................................................26 2.6.6 Post-hybridization......................................................................................................29 Chapter 3. Results..............................................................................................................32 3.1 Basal meristem developing process in Streptocarpus rexii...........................................32 3.1.1 Germination and development process of Streptocarpus rexii...................................32 3.1.2 Meristem activity........................................................................................................32 3.1.3 Basal meristem position..............................................................................................35 3.2 The homologs of gibberellin genes in S. rexii analysis.................................................37 3.2.1 Isolation of SrGA2ox and SrGA20ox..........................................................................37 3.2.2 Homology assessment of SrGA2ox and SrGA20ox....................................................37 3.2.2.1 Sequence identity......................................................................................................37 3.2.2.2 Phylogenetic tree construction of gibberellins genes..............................................38 3.2.2.3 Verification of SrGA2ox and SrGA20ox from conserved domain shared...............42 3.3 The temporal expression analysis of SrGA2ox and SrGA20ox by RT-PCR..................45 3.4 The spatial expression analysis of SrGA2ox and SrGA20ox by RNA in situ hybridization...................................................................................................................47 3.4.1 Gibberellin genes expressions in the basal meristem result in anisocotyly during seedling stages............................................................................................................47 3.4.2 Gibberellin genes expressions in embryo stages related to cotyledon determination.50 Chapter 4. Discussion.........................................................................................................53 4.1 SrGA2ox and SrGA20ox are homologs to other model species......................................53 4.2 SrGA2ox involved in basal meristem maintenance for anisocotyly formation...............53 4.3 SrGA2ox exhibited lateral dominance.............................................................................54 4.4 Macrocotyledon still needs a certain amount of gibberellin...........................................55 4.5 Cotyledon fate was established after germination but not determined in embryo stage.56 4.6 SrGA20ox expression in the embryoic shoot meristem correlation to the loss of apical dominance in Streptocarpus rexii...................................................................................57 4.7 The determined cotyledon fate is irreversible................................................................58 Chapter 5. Conclusion and Future Prospects..................................................................60 Chapter 6. Appendices........................................................................................................62 6.1 The metabolic pathway of gibberellin.............................................................................62 6.2 Typical and atypical results of surgical treatment in Streptocarpus rexii......................63 6.3 The proportion of the different growth patterns after the surgical treatment.................64 6.4 The remaining cotyledon area measurement at the different removal times of Streptocarpus rexii.........................................................................................................65 6.5 The differences in growth pattern between Streptocarpus rexii seedlings without an excised cotyledon versus those with excised cotyledon.................................................66 6.6 The different primer sites of the partial length sequence of SrGA2ox............................68 6.7 The different primer sites of the full length sequence of SrGA20ox..............................69 Chapter 7. References........................................................................................................70 Index of Figures and Tables Figures Fig. 1 Illustration of the phyllomorphic organization in Streptocarpus rexii........................9 Fig. 2 The gibberellin treatment in Streptocarpus rexii.........................................................9 Fig. 3 Model of interactions between KNOX protein, gibberellin genes and cytokinin gene in the shoot apical meristem.........................................................................................9 Fig. 4 The germination process in anisocotyly of Streptocarpus rexii.................................33 Fig. 5 Meristem activity detected by Aniline Blue staining.................................................34 Fig. 6 Meristem position revealed by DAPI staining...........................................................36 Fig. 7 Phylogenetic tree of SrGA2ox....................................................................................40 Fig. 8 Phylogenetic tree of SrGA20ox..................................................................................41 Fig. 9 Conserved region and sequence identity of SrGA2ox................................................43 Fig. 10 Conserved region and sequence identity of SrGA20ox............................................44 Fig. 11 RT-PCR revealed SrGA genes’ temporal expressions.............................................46 Fig. 12 Spatial localization of the gibberellin genes expressed in the developing seedling by RNA in situ hybridization........................................................................................48 Fig. 13 Spatial localization of the gibberellin genes expressed in the different embryo stages by RNA in situ hybridization...................................................................................51 Tables Table. 1 The list of primer pairs for different purposes regarding SrGA genes...................61 | |
| dc.language.iso | zh-TW | |
| dc.subject | GA20-oxidase | zh_TW |
| dc.subject | 子葉不等大 | zh_TW |
| dc.subject | Streptocarpus rexii | zh_TW |
| dc.subject | basal meristem | zh_TW |
| dc.subject | groove meristem | zh_TW |
| dc.subject | GA2-oxidase | zh_TW |
| dc.subject | Anisocotyly | en |
| dc.subject | GA20-oxidase | en |
| dc.subject | GA2-oxidase | en |
| dc.subject | basal meristem | en |
| dc.subject | Streptocarpus rexii | en |
| dc.subject | groove meristem | en |
| dc.title | 吉貝素在堇蘭不等大子葉發育中扮演的角色 | zh_TW |
| dc.title | The role of gibberellin metabolic genes in anisocotyly development of Streptocarpus rexii (Gesneriaceae) | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 趙淑妙,蔡文杰,陳淑華,陳仁治 | |
| dc.subject.keyword | 子葉不等大,Streptocarpus rexii,basal meristem,groove meristem,GA2-oxidase,GA20-oxidase, | zh_TW |
| dc.subject.keyword | Anisocotyly,Streptocarpus rexii,basal meristem,groove meristem,GA20-oxidase,GA2-oxidase, | en |
| dc.relation.page | 74 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-08-27 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生態學與演化生物學研究所 | zh_TW |
| 顯示於系所單位： | 生態學與演化生物學研究所 | |
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