碩士 / 大葉大學 / 分子生物科技學系碩士班 / 93 / Gonadotropins (GtHs), follicle stimulating hormone (FSH) and luteinizing hormone (LH) play a major regulatory role in reproductive process. They both contain a common glycoprotein α subunit (GtHα, common α) and a hormone-specific β subunit, such as FSHβ and LHβ, respectively. Zebrafish (Danio rerio) was used as a model fish and the technique of RNA interference (RNAi) was employed to knockdown these three glycoprotein subunits. Three short-hairpin RNA (shRNA) expression vectors and 3 miss-matched shRNA expression vectors as control for each subunit gene was constructed, and the depression efficiency was tested in vivo by microinjection and the RNA levels of GtHs gene were monitored by RT-PCR. The data revealed that the expression of GtHα and FSHβ mRNA was obviously depressed. The GtHs-GFP expression vectors for each subunit gene was further constructed, and changes of these three subunits at the protein level were stimulated by co-injection of GtHs-GFP expression vector and GtHs-shRNA expression vector in vivo. The data indicated that the shRNA knockdown efficiency is highly related to the forms of vector construction, and the best construction can knockdown GtHα subunit expression as much as 89.53%.This study also suppressed the GtHα gene expression by morpholino, the data indicated that GtHα morpholino led to the suppression of embryonic development and also embryonic mutant as the result of the injection of GtHα-shRNA. In the result of co-injection morpholino can knockdown GtHα subunit expression as much as 94.32%. However, attainment of fish infertility by shRNA expression vectors with considerations of tissue-specific promoters, remains to be further investigated.
| Identifer | oai:union.ndltd.org:TW/093DYU00061009 |
| Date | January 2005 |
| Creators | Jung-Chen Hsieh, 謝榮振 |
| Contributors | Jyh-Yih Chen, Wei-Tung Huang, 陳志毅, 黃尉東 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 150 |
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