Study On Alpha-amylase Gene5′Flanking Sequences From Ctenopharyngodon Idellus

Α-amylase is an importance enzyme to obtain energy for the biological. In theriver ecosystem, food chain system was constituted by the different feeding habits fish,access to energy in different ways to different fish. However, in the process of theevolution, the fish α-amylase gene sequence maintained the similarity. The studies ofthe α-amylase gene expressed sequence, is significance to reveal the mechanism ofthe functional groups of fish feeding.5′flanking sequence of grass carp alpha amylase gene, length of2,063bp, wasgotten by cloned with chromosome walking method. Using network bioinformaticsprediction software analyzing alpha amylase gene5′flanking sequences of grass carp,the predicted grass carp alpha amylase gene promoter regions is located at-252bp to-2bp; Grass carp alpha amylase gene transcription start site (TSS) was in theinitiation codon (ATG) upstream of30bp; Prediction of grass carp alpha amylasegene promoter regions include basic start of substructure components: TATA box andCAAT box. The analysis of transcription factor binding sites show that, grass carpalpha amylase gene5′flanks-385bp loci exist HNF3regulatory elements, and thesequence is:"5-TTTTTTTGT-3". In addition, there are5GR regulatory elements,including a palindrome structure, with the sequence as the "AGTTCTtgttgtTGTTCT",which may have a response induced by glucocorticoid.In the same way, alpha amylase gene5′flanking sequence of Yellowcheck carp,Mud carp and Bluntnose black bream were gotten by cloning, with the length of2,268bp,1,635bp and2,037bp, respectively. Grass carp, Yellowcheck carp, Mudcarp and Bluntnose black bream representative different freshwater bream feedingfishes of the Pearl River. The preliminary study found that the alpha amylase gene5′flanking sequence about240bp upstream area of the four cyprinids is highlyconserved, which shows that the start codon to about240bp upstream areas is thebasic function unit of controlling the gene expression.According to the results of the evolutionary tree constructed by alpha amylasegene5′flanking sequences, five species of fish as cypriniformes gathered for a class,perch gathered to a class and three copies of the gene for a puffer gathered for a class;It is showing that differentiation mechanism of alpha amylase gene5′flanking sequence of different predators between different family or order have differentpatterns.The system of evolutionary tree built by the part of alpha amylase gene exonreflects the evolution of the alpha amylase and kinship relations. Grass carp, zebrafishand mullet was gathered for a class, which belong to the Cyprinidae; the half-smoothtongue-sole and Winter flounder belonged to pleuronectiformes get together for aclass; the orange-spotted grouper, red eea bream, Bluefin Tuna, Barramundi andChinese perch get together for a class, which belong to perciformes; Chinese perchbelonged to freshwater fish in perciformes get together for a class and the rest of thefour kinds of saltwater fish together for a class. The evolutionary tree of results showthe fish alpha amylase gene sequence difference is consistent with the traditionalmorphological classification properties.The intron exon rule of alpha amylase gene in grass carp follows "GT-AG" rulesin this research, and the AT%of intron sequence base composition was obviouslyhigher than that of exons. The partial length of alpha amylase gene exon in grass carpis417bp, which can deduce that alpha amylase in the amino acid sequence containstwo alpha helix and three beta fold, and three forms the disulfide bond Cys site.