Howard Hughes Medical Institute, Department of Microbiology and Molecular Genetics, Davis, University of California, Davis, CA, USA.
Methods Mol Biol. 2021;2153:267-286. doi: 10.1007/978-1-0716-0644-5_19.
Crossing-over between homologous chromosomes is essential for accurate chromosome segregation at anaphase-I of meiosis. Defective crossing-over is associated with infertility, pregnancy miscarriage, and congenital disease. This chapter presents optimized protocols for the analysis of meiotic crossovers at the cytological level in spermatocytes and oocytes from mouse. The first approach employs immunocytology to detect MLH1, a DNA mismatch-repair protein that specifically marks crossover sites in the pachytene stage of meiotic prophase-I. These immunocytological methods have general utility for the analysis of other recombination steps, such as initiation and DNA strand exchange. The second approach visualizes chiasmata, the points of physical exchange between homologous chromosomes that are present during the diakinesis and metaphase-I stages. Both approaches are readily adaptable to the analysis of crossing over in other vertebrate species.
同源染色体之间的交叉互换对于减数分裂后期 I 的染色体正确分离至关重要。交叉互换缺陷与不孕、妊娠流产和先天性疾病有关。本章介绍了在小鼠精母细胞和卵母细胞的细胞学水平上分析减数分裂交叉的优化方案。第一种方法采用免疫细胞化学检测 MLH1，一种 DNA 错配修复蛋白，它特异性地标记得精前期 I 的粗线期的交叉位点。这些免疫细胞化学方法对于分析其他重组步骤，如起始和 DNA 链交换，具有普遍的适用性。第二种方法观察减数分裂前期 I 的终变期和中期的同源染色体之间发生物理交换的交叉点。这两种方法都可以很容易地适应于其他脊椎动物物种的交叉互换分析。
