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標題: | 甘薯SPORAMIN基因群的研究 Sporamin Gene Family of Sweet Potato |
作者: | 陳仁治 |
出版年 : | 1994 |
學位: | 碩士 |
摘要: | 以Sporamin反義基因SP-B cDNA為探針,對甘薯塊根cDNA基因庫進行篩選。從24個正反應株中得到了4個Sporamin cDNA(pGEM-TIA; pGEM-TIB; pGEM-TIC; pGEM-TID)。pGEM-TIA和pGEM-TIB包含了一個長660 bp的open reading frame可轉譯出一219個胺基酸,分子量24 kD,等電點5.8的蛋白質。完整的pGEM-TIA為0.9 kb;而pGEM-TIB為0.87 kb。pGEM-TIC和pGEM-TID則分別為0.65 kb及0.52 kb不完整的Sporamin cDNA。和已有的Sporamin cDNA(pIMO23、pIMO335、pIMO336及pIMO535)作比對,發現所得四條Sporamin cDNA和Sporamin A次基因群cDNA同源性為92-98%,和Sporamin B次基因群同源性為79-82%,因此所得四條cDNA皆屬Sporamin A次基因群。而pGEM-TIA和pIMO335除了具有不同的polyadenylation site外,為相同的cDNA。證實Sporamin cDNA具有Alternative polyadenylation site的現象。 取不同生長時期的塊根,觀察在不同時期Sporamin基因表現的情形。發現隨著塊根的生長Sporamin基因的表現越強,但塊根發芽後Sporamin基因便停止表現。取不同組織,觀察在不同組織Sporamin基因表現的情形,也發現,在莖部Sporamin基因有少量的表現,而葉部則不見Sporamin基因的表現,因此Sporamin為一甘薯塊根特定表現基因。 將Sporamin cDNA接在表現型載體(pGEX-2T)上,表現出Sporamin蛋白。發現全長的Sporamin先驅蛋白少掉N端18個或25個胺基酸的訊息多鏈,便具有trypsin inhibitor活性,全長的Sporamin先驅蛋白則沒有觀察到。此一trypsin inhibitor活性推測和植物防禦有關。 Four sporamin cDNAs were isolated from sweet potato tuberous root cDNA library with 32P-labelled cDNA inser of SP-B, a sporamin antisense cDNA clone, as a hybridization probe. These four cDNA clones, pGEM-TIA, pGEM-TIB, pGEM-TIC, pGEM-TID, carry cDNA insers of 0.9, 0.87, 0.65, 0.52 kb respectively. The pGEM-TIA and pGEM-TIB carry full-length cDNAs for sporamin mRNA. Their cDNAs clones share 92-98% identity with spoamin A (pIMO23, pIMO335, gSPOA1) and 79-82% idntity with sporamin B (pIMO336, pIMO535, gSPOB1). This result suggests theses cDNA clons belong to sporamin A gene subfamily. pIMO335 has same coding region and 5' noncoding region with pGEM-TIA, but an erly polyadenylation site is found in the 3' noncoding region of pIMO335. This fact suggests possible alternal polyadenylation phenomenum in sporamin genes. Sweet potato tuberous roots of different size were used to investigate the sporamin genes expression. As tuberous root enlarge, the sporamin mRNA accumulate to a higher level, and reduced to nondetectable white the tuberousroot sprouts. The PCR-amplifed-coding region of pGEM-TIA were subcloned into pGEX-2T to examine whether sporamin is a typsin inhibitor. The exprassion product lacking 18 or 25 amino acid residues in the N-terminal show trypsin inhibitor activity, but no observable activity was found in full-length expressed sporamin protrins. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/76053 |
全文授權: | 未授權 |
顯示於系所單位: | 植物科學研究所 |
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