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目的研究五氯酚钠(NaPCP)对斑马鱼体细胞p53基因的致突变作用。方法设置5个不同浓度的NaPCP暴露组及空白对照组,采用Tubingen品系斑马鱼进行斑马鱼急性毒性试验计算半数致死浓度(LC50),并依据急性毒性实验所得结果,设置1/10 LC50暴露剂量,暴露组与空白对照组各取10尾斑马鱼,在连续暴露45 d后提取斑马鱼肝脏组织RNA,采用反转录-聚合酶链式反应扩增p53基因c DNA序列,并进行测序分析。结果NaPCP急性毒性实验所得LC50为18.4μg/L,测序分析结果显示斑马鱼p53基因编码区序列长1125 bp,编码374个氨基酸,与Gen Bank已知Tubingen品系斑马鱼p53序列(GI:425876786)同源性为99.2%,与其他生物物种的p53序列存在一定差异。在NaPCP 1.8μg/L浓度下暴露45 d后,与对照组比较,暴露组斑马鱼p53基因序列的8、148和229位密码子发生GAG→AAG、CAT→CAG和CAG→CAA突变,导致所编码的p53蛋白发生Glu→Lys和His→Gln变异。结论 NaPCP是一种致基因突变物,可诱导斑马鱼体细胞p53基因发生突变,对人体可能有潜在的致基因突变风险。
Objective To study the mutagenic effects of sodium pentachlorophenol (NaPCP) on p53 gene in zebrafish somatic cells. Methods Five different concentrations of NaPCP exposure group and blank control group were set up. The zebrafish (Tubingen strain) was used for zebrafish acute toxicity test to calculate LC50. According to the results of acute toxicity test, 1/10 LC50 exposure dose was set, The zebrafish liver RNA was extracted from the zebrafish liver tissue after exposed for 45 days. The p53 gene cDNA sequence was amplified by reverse transcription - polymerase chain reaction and sequenced. Results The LC50 of NaPCP acute toxicity test was 18.4 μg / L. Sequencing analysis showed that the coding region of the p53 gene of zebrafish was 1125 bp in length and encoded a protein of 374 amino acids. This sequence was identical to the p53 gene sequence of Tubingen strain (GI: 425876786) 99.2% of the origin, there are some differences with the p53 sequence of other biological species. After exposure to NaPCP 1.8μg / L for 45 days, GAG → AAG, CAT → CAG and CAG → CAA mutations occurred in codon 8,148 and 229 of the p53 gene of zebrafish exposed to the control group The encoded p53 protein undergoes Glu → Lys and His → Gln mutations. Conclusion NaPCP is a mutagenic gene that can induce the mutation of p53 gene in zebrafish somatic cells and may have potential risk of mutagenesis to human.