Several Wild Mouse 4.5s Gene Cloning And Sequence Analysis

The retroposon is a transposable element which can transpose among genome with the path of DNA — RNA — DNA. That is to say, it has the ability to be integrated into the host genome by an enzyme, so-called integrase. In this means, the host genome becomes very huge as the number of the retroposon copy increases. The retroposon often has regulator elements, such as enhancer, promoter, etc .These elements will influence the expression of the host gene. So we can say that the retroposon plays an important role in the evolution of life.There are different types of retroposon in mammalian genomes, such as LINEs, SINEs and pseudo genes, etc. LINEs and pseudo genes are transcribed by RNase II. SINEs are nonautonomous elements and transcribed by RNase III. SINEs can be divided into two groups according to their origins, ones come from tRNA, and others come from 7SL RNA. The former count the most, the latter include the B1 family, Alu family, as so on.The 4.5S RNA gene sequence which comes from 7SL RNA is a kind of retroposon that was discovered very late. It has its own transcription which is transcribed by RNase III and called 4.5S RNA. 4.5S RNA is one of the snRNAs. Small molecular RNAs, such as snRNA, snoRNA, siRNA and miRNA have become the focus in the area of life science.4.5S RNA was first isolated from some RNA virus in late 1960s'. Later it was discovered in rat, mouse and Chinese hamster cells that were not infected by virus. But it has not yet been discovered in human, domestic cat, rabbit and the chicken cells. Harada F et al (1980) analyzed the sequences of mouse and Chinese hamster 4,5S RNA and put forward the possible secondary structure. Not so long from that, 4.5S RNA was found associated with poly (A) RNA, which means that 4.5S RNA has something to do with hnRNA processing after hnRNA was transcribed. Perhaps it joins directly splicesome as a composition. Li (1982)found that 4.5S DNA has a gene sequence highly homologous with the gene sequences of 7S RNA, Bl and Alu. It may be one of the members of Bl family. Researches till today show that retroposons like Bl, Alu, perform many important functions, for example, regulating gene expression or participating in genomic evolution in cell. Other reports show there is an abundant small RNA called 4.5S RNA, which has a gene sequence homologous with that of 7S RNA of eukaryotes. Some experiments show when the 4.5S RNA is absent translation and secretion is effected. The research made by professor Xu lai-xiang (2000) show the relation between the 4.5S retroposon and enhancer in structure and functions. These facts tell us the 4.5S retroposon does carry out some important biological functions. But the research work up to today is still limited in a few species such as rat, mouse and Chinese hamster. The 4.5S retroposon of other species have not been researched yet.In this paper, the cDNA of 4.5S RNA was amplified by reverse transcription -polymerase chain reaction (RT-PCR) with total RNA extracted from Cricetulus triton de Winton, A.agrarius and C.barabensis liver tissues as the template for the first time. Then the cDNA gene was cloned into pUC19 plasmid and sequenced. These different sequences were analyzed and compared with each other. In this way, we found that the 4.5S retroposon gene sequence has so much GC in it which counts to be 60%. There are two repeat sequences in 4.5S DNA .One is G-C-C-G-G-T-A-G located in 1-8 site and 20-27 site, the other is G-A-G-G-C-A-G-A-G-G located in 40-49 site and 46-55 site. The fact that 4.5S retroposon is very conservative because there are a few different nucleotides in the 88 base pair which has not apparent influence on its secondary structure ,shows that it perform so important functions that it still remained after a long period of evolution.4.5S retroposon is mainly found in rodent species but few is found in other mammalian. This fact probably show on which aspect rodent species is special. The question about whether it exists in all rodent species ,how significant it is torodent species, what biological function on earth it performs, what role it plays in evolution of rodent genome, can only be answered after the further research on the structure and functions of 4.5S retroposon.