Abstract：
Across eukaryotes, genome stability is essential for normal cell function, physiology, and species survival. Aberrant expression of key genes or exposure to genotoxic agents can have detrimental effects on genome stability and contribute to the development of various diseases, including cancer. Chromosome instability (CIN), or ongoing changes in chromosome complements, is a frequent form of genome instability observed in cancer and is a driver of genetic and cell-to-cell heterogeneity that can be rapidly detected and quantitatively assessed using surrogate markers of CIN. For example, single cell quantitative imaging microscopy (QuantIM) can be used to simultaneously identify changes in nuclear areas and micronucleus formation. While changes in nuclear areas are often associated with large-scale changes in chromosome complements (i.e., ploidy), micronuclei are small extra-nuclear bodies found outside the primary nucleus that have previously been employed as a measure of genotoxicity of test compounds. Here, we present a facile QuantIM approach that allows for the rapid assessment and quantification of CIN associated phenotypes and genotoxicity. First, we provide protocols to optimize and execute CIN and genotoxicity assays. Secondly, we present the critical imaging settings, optimization steps, downstream statistical analyses, and data visualization strategies employed to obtain high quality and robust data. These approaches can be easily applied to assess the prevalence of CIN associated phenotypes and genotoxic stress for a myriad of experimental and clinical contexts ranging from direct tests to large-scale screens of various genetic contexts (i.e., aberrant gene expression) or chemical compounds. In summary, this QuantIM approach facilitates the identification of novel CIN genes and/or genotoxic agents that will provide greater insight into the aberrant genes and pathways underlying CIN and genotoxicity.
摘要：
在真核生物中，基因组稳定性对正常细胞功能至关重要，生理学,物种生存。关键基因的异常表达或暴露于基因毒性因子会对基因组稳定性产生有害影响，并导致各种疾病的发展。包括癌症.染色体不稳定性（CIN），或者染色体互补的持续变化，是在癌症中观察到的基因组不稳定的常见形式，并且是遗传和细胞间异质性的驱动因素，可以使用CIN的替代标记进行快速检测和定量评估。例如,单细胞定量成像显微镜（QuantIM）可用于同时识别核区和微核形成的变化。虽然核区的变化通常与染色体互补序列的大规模变化有关(即，倍性)，微核是在初级核外发现的小核外体，以前曾被用作测试化合物遗传毒性的量度。这里,我们提出了一种简便的QuantIM方法，可以快速评估和定量CIN相关表型和遗传毒性。首先,我们提供了优化和执行CIN和遗传毒性测定的方案。其次,我们提出了关键的成像设置，优化步骤,下游统计分析，以及用于获取高质量和健壮数据的数据可视化策略。这些方法可以很容易地应用于评估CIN相关表型和遗传毒性应激的患病率，包括从直接测试到各种遗传背景的大规模筛选的无数实验和临床背景（即，异常基因表达）或化合物。总之,这种QuantIM方法有助于鉴定novelCIN基因和/或遗传毒性因子，这将更深入地了解CIN和遗传毒性的异常基因和通路。
