Construction and identification of differential expression genes of peripheral blood cells in radon-exposed mice

Abstract

Objective To screen and identify the differential expression genes on peripheral blood cells of mice based on the experimental animal model of radon exposure. Methods BALB/c mice were exposed in a type HD-3 multifunctional radon-room, with the accumulative doses of radon-exposure group at 105 WLM and control group at 1 WLM. Total RNA was extracted from peripheral blood cells and the methods of SMART for dscDNA synthesis and SSH for gene screening was applied. With the construction of the cDNA library enriched with differentially expressed genes, the pMD 18-T plasmid containing LacZ operator at the multiple cloning site was used to allow a blue-white screening. The TA clones were amplified by nested PCR and the reverse Northern blot was used to identify up and down regulation of the clones. The differently expressed cDNA was then sequenced and analyzed. Results The subtracted cDNA libraries were successfully constructed. A total of 390 recombinant white colonies were randomly selected. Among the 312 cDNA monoelones selected from bath forward- and reverse-subtracted libraries,41 clones were chosen to sequence for their differential expressions based on reverse Northern blot. Among the 41 sequenced clones, 10 clones with known function/annotation and 3 new ESTs with the GenBank accession numbers were obtained. Most of the known function/annotation genes were revealed to be related with cell proliferation, metabolism, cellular apoptosis and carcinogenesis. Conclusions The animal model of radon exposure was established and the cDNA library of peripheral blood cells was suceessfully constructed. Radon exposure could up- and down-regulate a series of genes. Differentially expressed genes could be identified by using SSH technique and the results may help exploring mechanisms of random exposure. Key words: Radon; Suppression subtractive hybridization; Differential expression genes; Bioinformatics