Chang Zhong-Jie, Chen Ying, Du Qi-Yan
College of Life Sciences, Henan Normal University, Xinxiang 453002, China.
Yi Chuan Xue Bao. 2003 Jun;30(6):528-32.
Oxygen content in water is one of the limit factors to the fish growing. The tolerance of fish to low oxygen content depends on the affinity of hemoglobin to oxygen. The tempt of transgenic fish study is to transfer hemoglobin gene from the fish that tolerates to low oxygen content in water to the fish sensitive to low oxygen condition. Therefore, using hemoglobin gene to breed new varieties may become a main trend in the near future. To explore the underline mechanisms for the variation of hemoglobin property, we have cloned hemoglobin-alpha-chain from five species of Cypriniformes fishes. A pair of consensus degenerate primes were designed basing on N-terminal and C-terminal conservative amino acid of alpha-chain. Using RT-PCR method, alpha-globin gene was amplified and cloned from total RNA which was extracted from blood of five fishes which are widely distribution in China (Cyprinus carpio, Ctenopharyngodon idellus, Carassius auratus, Misgurnus anguillicaudatus, Paramisgurnus dabryanus). Sequence comparison with the existing hemoglobin alpha-chain confirmed that these cDNA fragments were alpha-globin gene of five fishes. The accession numbers in GenBank are AF528156, AF528157, AF528197, AF528198, AF528199 respectively. The cDNA which coding amino acid are all 429 bp. Compared with the amino acid sequences of five fishes, it can be concluded that in five fishes the highest similarity of two fishes which are Paramisgurnus dabryanus and Misgurnus anguillicaudatus is 99.41%, whereas the lowest similarity of two fishes which are Ctenopharyngodon idellus and Paramisgurnus dabryanus is 83.92%. In addition, the sequence similarity and phylogenetic relationship were compared with Within-group method. The result shows that there is much consistent between similarity and phylogenetic relationships from morphological of those near relation species and much connection between similarity and environment of those distant relation species.
水中的氧气含量是鱼类生长的限制因素之一。鱼类对低氧含量的耐受性取决于血红蛋白对氧气的亲和力。转基因鱼研究的诱人之处在于将耐低氧含量的鱼类的血红蛋白基因转移到对低氧条件敏感的鱼类中。因此，利用血红蛋白基因培育新品种可能在不久的将来成为一个主要趋势。为了探索血红蛋白特性变异的潜在机制，我们从五种鲤形目鱼类中克隆了血红蛋白α链。基于α链的N端和C端保守氨基酸设计了一对共有简并引物。采用RT-PCR方法，从中国广泛分布的五种鱼类（鲤鱼、草鱼、鲫鱼、泥鳅、大鳞副泥鳅）血液中提取的总RNA中扩增并克隆了α-珠蛋白基因。与现有的血红蛋白α链进行序列比较，证实这些cDNA片段是五种鱼类的α-珠蛋白基因。在GenBank中的登录号分别为AF528156、AF528157、AF528197、AF528198、AF528199。编码氨基酸的cDNA均为429bp。通过对五种鱼类氨基酸序列的比较可以得出，在五种鱼类中，大鳞副泥鳅和泥鳅的相似度最高，为99.41%，而草鱼和大鳞副泥鳅的相似度最低，为83.92%。此外，采用组内法比较了序列相似性和系统发育关系。结果表明，近缘物种形态学上的相似性与系统发育关系之间有很多一致性，远缘物种的相似性与环境之间有很多联系。
