Primary Studies On The Cloning Of The Cross-intron Genomic DNA Sequences And The Alternative Splice Of Tra-1 Genes From C.elegans

The control of eukaryotic gene expression is currently a key focal point of life science research. Research has shown that mRNA affects gene regulation and expression, so splicing, especially alternative splicing of RNA, will affect the control of eukaryotic gene expression. Eukaryotic pre-mRNAs are spliced to form mature mRNA. The pre-mRNAs alternative splicing greatly increases the diversity of proteins and the complexity of genes expression. Although the mechanism of alternative splice is unknown, these sequences must have importance biology function. So analysis of intron in genes has important sense for moleculemechanism of gene transcript and function.Tra-1 gene is an important gene in C.elegans, which is one of the most important model organisms. And tra-1 gene is the terminal control gene for somatic sex determination in C.elegans. Tra-1 gene encodes five zinc finger protein, in the second and third zinc finger of tra-1 gene the alternative splicing happens, so the gene has two transcript: one is an 1.5kb transcript that encodes a protein with two zinc finger motifs; and the other is a 5kb transcript that encodes a protein with five zinc fingers. The different outcome protein activity of tra-1 gene depends on the alternative splice of mRNA processing, which governs the number of zinc fingers in the resulting TRA-1 protein.In this paper, the upstream and downstream primers were designed in the first and fifth zinc finger respectively, and the 3.3kb PCR product had been obtained from the C.elegans genome DNA, then the PCR product had been cloned into pBS-T vector, which can transform E.coli TG1 and got more proliferation. 3.3kb PCR product still obtained from the recombined E.coli TG1, testified that the recombined plasmid succeeds. Comparison of the genome DNA and cDNA sequence revealed that the distal polyA site of 1.5kb short mRNA was the same as one intron , whose secondary structure showed that the polyA sites in the active point, but the site of the 5' splicing signal GU was not sure to activity. So thealternative splicing of tra-1 mRNA maybe happened.The whole procedure initiates a novelty technique in the study of Mermithidae. What is more, it has settled a rigid foundation for the further research of the gene in sex detennination.