编码四种甲壳动物蜕皮抑制激素的部分cDNA的克隆和序列分析

IOCAS-IR > 海洋环流与波动重点实验室

	编码四种甲壳动物蜕皮抑制激素的部分cDNA的克隆和序列分析
	王在照
学位类型	博士
	2000
学位授予单位	中国科学院海洋研究所
学位授予地点	中国科学院海洋研究所
学位专业	海洋生物学
关键词	中国对虾中华绒螯蟹鹰爪虾蓝对虾蜕皮抑制激素 Cdna克隆序列分析
摘要	本文分别提取了中国对虾、中华绒螯蟹、鹰爪虾和蓝对虾的眼柄总RNA。用无DNA污染的眼柄总RNA为模板，根据日本对虾的MIH设计兼并引物P1和P2，进行RT－PCR扩增，在适宜的反应条件下，均分别得到一特异性的产物。以蓝对虾基因组DNA为模板的PCR扩增也能得到一与特异性cDNA片段大小相似的特异性的DNA片段。分别将这些片段亚克隆到载体中进行测序，测得中国对虾特异性cDNA片段由203个碱基组成，中华绒螯蟹、鹰爪虾、蓝对虾的特异性cDNA片段及蓝对虾的特异性DNA片段由215个碱基组成，其中蓝对虾特异性cDNA与由蓝对虾的基因组DNA扩增得到的特异性DNA片段的碱基序列几乎完全相同。利用互联网上的在线工具Fasta3查找这些cDNA推定的氨基酸序列的相似序列，可以发现大量的甲壳动物的CHH家族的神经肽与它们相似。在所有的相似序列中，分别由中国对虾、中华绒螯蟹、鹰爪虾和蓝对虾的特异性cDNA片段推定的氨基酸序列与蜕皮抑制激素（MIH）的相似度最高，这一结果提示分别由中国对虾、中华绒螯蟹、鹰爪虾和蓝对虾的眼柄特异性cDNA片段推定的氨基酸序列可能是这四种甲壳动物的MIH片段。根据核苷酸数据和氨基酸数据，比较这四种甲壳动物的MIH以及已发表的六种MIH之间的相似度，同时分析这十种MIH的系统进化关系，结果是蓝对虾、鹰爪虾和中华绒螯蟹的MIH相似性较高，亲缘关系较近，这与物种的系统演化不相符。
其他摘要	Total RNAs were extracted from eyestalks of the shrimps (including Penaeus chinensis, Trachypenaeus Curvirostris, and Penaeus stylirostris) and the crab Eriocheir sinensis. Eyestalk complementary DNAs were gotten from these DNA-free total RNAs by reverse transcription. Based on the amino acid sequence of the molt-inhibiting hormone from Penaeus japonicus, two primers P_1 and P_2 were designed. Polymerase Chain Reaction (PCR) was performed using P_1 and P_2 as primers and these eyestalk cDNAs as templates. Four specific cDNAs were cloned under proper reaction conditions. At the same time in a control of PCR whose template is genomic DNA of P. stylirostris, a specific DNA fragment of the same size as the specific cDNA from eyestalk of P. stylirostris was cloned. These specific cDNAs and DNA were subcloned into vector for sequencing. The sequencing results demonstrated that the specific cDNA from P. chinensis eyestalk consists of 203 base pairs, and all the specific cDNAs from the shrimps T. curvirostris, P. stylirostris and the crab E. sinensis consist of 215 base pairs. The specific DNA from genomic DNA of the shrimp P. stylirostris is almost as same as the specific cDNA from P. stylirostris eyestalk. Using the on-line sequence similarity searching tool Fasta3 on interned web, many neuropeptides from crustacean CHH family of high similarity with these deduced amino acid sequences from cDNA of eyestalks from these four kinds animals were found. The alignments of the deduced amino acid sequences from eyestalk cDNAs of the shrimps P. chinensis. T. curvirostris, and P. stylirostris and the crab E. sinensis with neuropeptides of CHH family from various crustaceans demonstrate that they have the highest identity and homology with most MIHs from crustaceans. It suggests that the deduced amino acid sequences from eyestalk cDNA of the shrimps P. chinensis, T. curvirostris, and P. stylirostris and the crab E. sinensis are the part of MIH neuropeptides of corresponding animals. By using nucleotide acid sequences and amino acid sequences data, The relationship among MIHs from P. chinensis, E. sinensis, T. curvirostris, and P. stylirostris and six published MIHs from various crustacean was analyzed. The gene tree of MIH disagrees with phylogenetic tree of species. The relationship among E. sinensis. T. curvirostris, and P. stylirostris is very near.
页数	124
语种	中文
文献类型	学位论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/361
专题	海洋环流与波动重点实验室
推荐引用方式 GB/T 7714	王在照. 编码四种甲壳动物蜕皮抑制激素的部分cDNA的克隆和序列分析[D]. 中国科学院海洋研究所. 中国科学院海洋研究所,2000.

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