生物样品体外致突变高通量检测方法研究进展

Advance in Researches on High-throughput Methods for Mutagenicity Evaluation of Biological Samples In Vitro

评述了几种常用的体外致突变检测方法及其用于生物样品检测的可行性,有些方法经改进可用于高通量检测生物样品体外致突变性.经典Ames实验受生物样品中组氨酸的影响,易产生假阳性结果,尽管经过修正可以排除组氨酸的干扰,但操作繁琐,不适合高通量检测.基于SOS反应的检测体系避开了组氨酸的影响,且简单易行,适合高通量检测:以β-半乳糖苷酶基因(lacZ)作为报告基因的检测体系灵敏度高,且经过离心洗涤或后培养的方式可降低样品颜色的影响;以绿色荧光蛋白(GFP)基因作为报告基因的检测体系避开了颜色的干扰,但这类方法灵敏度普遍不高,可以寻找信号更强的荧光蛋白以替代GFP;荧光素酶(lux)基因集lacZ和GFP的优点于一身,但检测时需要额外添加辅助因子,限制了其应用.也对单细胞凝胶电泳、tk基因突变实验、染色体损伤检测等方法进行了分析,有些适合生物样品高通量检测,但由于缺少国际通用的标准,很难推广使用.

Several mutagenicity test methods and their adaptability in safety evaluation of biological samples are presented and discussed in this paper.After modification,some of these assays can be used in high-throughput test of biological samples.Classical Ames-test is not adaptable to detect the mutagenicity of biological samples containing histidine or proteins.Although false positive results can be avoided after modification,Ames-test can’t be applied in a high-throughput way for its complicated procedures.Methods based on SOS response can avoid the influence of histidine or proteins.Most of them can be used for highthroughput assessment of the in vitro mutagenicity of biological samples.Colorimetric assay is used in umu-lacZ test whose reporter is β-galactosidase.The interference of colors of biological samples can be eliminated by either centrifugal washing or post-incubation.Systems with GFP as reporters avoid the interference of colors of biological samples.Although SOS/GFP tests is not sensitive,more robust reporters can be sought to substitute GFP.Luciferase has both advantages of β-galactosidase and GFP,but it is not used widely because its reaction needs additional cofactors.In addition,single cell gel electrophoresis assay,thymidine kinase gene mutation assay,chromosome damage assay such as micronucleus test,sister chromatid exchange test and chromosome aberration assay were analyzed and compared.Some of these systems are adaptable to assess the in vitro mutagenicity of biological samples,but they won’t be applied extensively because there are no international universal standards about these detection systems.