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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/34599

Title: Tbx5基因缺損對斑馬魚(Danio rerio)胸鰭軟骨生成之影響
The impact of tbx5 gene knockdown on pectoral fin and chondroginesis during zebrafish (Danio rerio) embryonic development
Authors: Jia-He Wang
王佳禾
Contributors: NTOU:Department of Aquaculture
國立臺灣海洋大學:水產養殖學系
Keywords: tbx5;cdh2;軟骨生成
tbx5;cdh2;chondrogenesis
Date: 2013
Issue Date: 2013-10-07T02:42:57Z
Abstract: Tbx5屬於T-box家族中的轉錄因子,在脊椎動物胚胎發育時期表現於心臟、前肢以及眼睛。本實驗利用專一反意股寡核甘酸抑制斑馬魚胚胎tbx5基因表現,在37hpf的斑馬魚胸鰭上會造成軟骨凝結停止的現象。以斑馬魚DNA晶片偵測tbx5基因弱化後斑馬魚胚胎在24、30、37及48hpf基因表現情形,在1.5倍基因表現量下降的基因並利用軟體篩選出與軟骨凝結相關的基因cdh2。利用晶片的結果來進行相對即時定量偵測軟骨凝結相關基因cdh2在斑馬魚受精後26、30、37以及48小時之表現,4個時間點在tbx5基因弱化後期表現量皆有顯著下降(p<0.05)。在免疫螢光染色的實驗結果顯示軟骨凝結蛋白CDH2在tbx5-MO處理後在37hpf、48hpf皆有下降,推測tbx5與軟骨凝結有直接或間接的關係。利用生長相關荷爾蒙GH、IGF-1改善tbx5基因所造成的缺損,在相對即時定量的結果中cdh2基因經生長荷爾蒙GH、IGF-1改善後表現皆有上升。在免疫螢光染色實驗中,在tbx5-MO添加荷爾蒙GH、IGF-1後CDH2蛋白表現量皆有上升。由以上結果可以推測當tbx5基因弱化後會導致與軟骨凝結及細胞黏附相關的cdh2表現量下降,進而影響到斑馬魚胸鰭軟骨的發育。
Tbx5 is a member of the T-box transcription factor family which expresses in the heart for limbs/pectoral fin and eyes of vertebrate animals during embryonic development. In zebrafish embryos, using the specific antisense oligonucleotide to inhibit the tbx5 gene expression, the cartilage condensation of pectoral fin ceases. The DNA microarray revealed the expression of cartilage development related genes declined in tbx5 gene knockdown embryos, further analyzed and selected the cell adhesion and cartilage condensation gene cdh2. Furthermore, Real-time PCR analysis indicated that mRNA expression of cdh2 descend at 26, 30, 37 and 48 hpf. Immunohistochemistry staining results indicated that the protein level expression of CDH2 declined in tbx5 knockdown treatment. In present study, Real-time PCR analysis indicated that mRNA expression of cdh2 in co-injection of tbx5-MO and GH or tbx5-MO and IGF-Ⅰ zebrafish embryos increased . Immunohistochemistry staining results indicated that the protein level expression of CDH2 increased in tbx5 knockdown treatment.
URI: http://ethesys.lib.ntou.edu.tw/cdrfb3/record/#G0010033008
http://ntour.ntou.edu.tw/handle/987654321/34599
Appears in Collections:[水產養殖學系] 博碩士論文

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