The Studies On Cloning And Expression Of QingDao Amphioxus, Sox1/2/3-like Genes

Sox gene family is composed of SRY (region of sex determination Y chromosome) genes related to sex determination. They are characterized by the presence of a HMG domain. It can widely participate in the early embryonic development and neuro development in the process of the individual development. Researching the functions and features of the Sox gene family is of great significance for revealing the molecular mechanism of sex determination, gene regulation for embryonic development and understanding the development process of various tissues and organs.Amphioxus belong to cephalochordate, which is the existent closest relative groupes between invertebrate and vertebrates, since it has part of the morphological pattern of the vertebrates gene multiplied not so great and not too complex, is become the model organisms in the study of origins and evolution of the vertebrates.The earlist homology Sox gene which is found in amphioxus is Amphisox1/2/3 gene. The expression of this gene in Branchiostoma floridae is closely related with the occurrence of outside nerve mesoderm and the formation of the neural tube. In 2005, Zhangwei also found homology Sox in Branchiostoma belcheri tsingtauense, and named Amphisox1/2/3-like genes.(GeneBank number:AY870807). When the endoderm of the embryos began differentiate, this gene expressed in the original bowel. Alone with the development of embryo, its expression regional successively appeared in neural board, neural tube and accord with the development of the central nevous system. At the same time, along with the completed of the central nervous system development, its expression level gradually lowered from formerly to backward. The space-time expression patterns of Amphisox1/2/3 suggests that it closely related with the happening of nevous system in Branchiostoma belcheri tsingtauense .The study of comparing with the development relative genes of amphioxus and vertebrate reveals that in evolutionary,the genetics mechanism of the outside nerve mesoderm and not outside nerve mosederm differentiation appeared in the period before amphioxus and vertebrate begin differentiating. Sox, the level of the expression network of this gene in outside nerve mesoderm is still not clear, to begin the studies of Amphisox1/2/3-like genes in Branchiostoma belcheri tsingtauense is very significant.The present article through seeking from GeneBank with the Sox genes sequences those sibship closest with amphioxus Sox genes such as zebra fish, the handle ascidia and so on. Designing a pair of specificity primer according to these conservative Sox gnens sequence and using PCR method, we get the segment of Sox1/2/3-like gene in Branchiostoma belcheri tsingtauense. According to the gene segment which has been already obtained, using genome walking methord to get 3 end and 5 end unknown flanking sequence. Using PCR method and labeling the Digoxin on the probe, with amphioxus tissue slice, the expression regional of this gene in Branchiostoma belcheri tsingtauense is examined by in situ hybridization. In addition, in the use of the characteristic of antigen specified united with the immune body in the technology of electron microscospic in situ hybridization examined and located expression regional of the Sox1/2/3-like gene in Branchiostoma belcheri tsingtauense cells.