Study On The Expression Of P53,p21 In Rat Liver During Tumor Promotion By Microcystin-LR And Related Mechanism

【Objective】This study was designed to determine the expression of p53,p21waf1 and the point mutation of p53 gene in rat liver during tumor promotion by MC-LR in order to explore the related molecular mechanism.【Motheds】1. The animal model was set up with two-stage-medium-term theory, DEN was applied as initiator and MC-LR applied as promoter. Rats were divided into three groups: blank control group, DEN control group, and MC-LR tumor promoting group.2. Expression of the p53, p21waf1 genes in rat liver were detected by RT-PCR (reverse transcriptase-polymerase chain reaction): Total RNA was prepared from the liver tissue by standard methods. The levels of p53 and p21 were semi-quantified by RT-PCR with endogenous internal standard, GAPDH. The PCR products were analyzed on a 2% agarose gel by electrophoresis. Gels were stained with ethidium bromide, visualized with UV transillumination, photographed, and submitted to image analysis. Quantitative image analysis of the PCR fragments was performed using the genesnap Image program. The mRNA levels were calculated as the ratios of optical density of the PCR products to those of the GAPDH PCR product.3. p53 gene mutations induced by MC-LR during rat two-stage hepatocarcinogenesis were screened by single-strand conformation polymorphism (SSCP) analysis, which is a convenient and rapid method for...