The CDNA Cloning And Expression Analysis Of Anxa-1、Anxa-2 Gene From Sika Deer Antler Tissue (Cervus Nippon Hortulorum)

The antlers are male secondary sex characteristic of cervidae, and they are the only mammalian appendages capable of regeneration. In rapid growing stage cell proliferation rate of tip tissue is nearly 30 times division rate of the tumor cells, and any mammalian tissues or organs can not compare with the fast growing rate. During the development, cell differentiation, proliferation and apoptosis were regulated under gene expression, but there still not any study could clear the antler growing mechanism fundamentally.The mesenchymal tissue of Northeast Sika Deer was used as experimental materials, and Annexins were taken as the research object, which had important connections with the tumor cells. The cDNA sequences of Anxa-1 and Anxa-2 genes were successfully cloned by RT-PCR technology. Bioinformatics method and Real-time Fluorescence Quantitative PCR technique were used to analyze the amino acid sequence and the expression at different phases. The main results were as follows:1 cDNA cloning and bioinformatics analyse of Anxa-1 and Anxa-2 genes from Northeast Sika Deer AnlterThe sequencing results showed that Anxa-1 and Anxa-2 genes were 1059bp and 1180bp respectively, and encoded 346 amino acids and 339 amino acids. Bioinformatics analysis showed that Anxa-1 and Anxa-2 proteins were both stable protein. Molecular weight were 38830.4 and 38612.0. The theoretical isoelectric point were 6.17 and 6.92. Leucine accounted for the largest proportion was 10.4% in the primary structure of Anxa-1 protein, and Leucine and Lysine in the primary structure of Anxa-2 protein accounted for the largest proportion were both 10.0%. Anxa-1 and Anxa-2 proteins had no signal peptide. The subcellular localization analysis showed that Anxa-1 protein was in the cytoplasm and Anxa-2 protein was in the nucleus. By predicting the secondary structure, we found that spatial structures of the proteins were both composed of alpha helix, random coil and extended chain, and the thirdary structure of the two proteins were constructed successfully. Multiple sequence alignment of the two proteins was done, and we found that these two genes conserved highly among different species, which suggested that the function was also similar in different species. By constructing the phylogenetic tree, we found that the two genes in the evolution were both divided into two branches, one branch was mammals, and the other was birds. Obviously that these two genes were evolved from a common ancestor gene, and both were important functional genes.2 Real-time Fluorescence Quantitative analysis of Anxa-1 and Anxa-2 genes from antler in different growth periodsReal-time Fluorescence Quantitative PCR results showed that the two genes expressed differently in different growth periods of antler. The expression level of Anxa-1 gene was higher at its earlier stage than at its intermediate and advanced stages, the expression level of Anxa-2 gene was higher at its intermediate stage than its earlier and advanced stages.