Evaluation of Crossover Number, Distribution, and Interference Using Cytological Assays in Arabidopsis
Evaluation of Crossover Number, Distribution, and Interference Using Cytological Assays in Arabidopsis
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使用拟南芥细胞学分析评估交叉数、分布和干扰
DOI：
10.1002/cpz1.599
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发表时间：
2022-12-01
期刊：
影响因子：
--
通讯作者：
Wang，Yingxiang
中科院分区：
文献类型：
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作者：
Huang，Jiyue;Li，Xiang;Wang，Yingxiang
减数分裂涉及亲本细胞中核染色体的复制，随后是两次连续的核分裂以产生单倍体孢子，其分化成配子体世代，其在植物中产生卵子和精子。减数分裂重组或交叉（CO）是减数分裂的标志，它允许同源染色体（同源物）之间的遗传信息改组，从而产生遗传多样性的后代细胞，并最终产生后代中的个体;这为遗传分化和物种形成提供了巨大的机会。减数分裂CO也确保二价体的形成和它们的平等分离的保真度。因此，调节减数分裂重组的机制已经在多个物种中被广泛研究。已经开发了几种方法来观察或估计CO的频率，其中CO可以通过估计交叉（复数交叉）的数量进行可视化和细胞学分析，交叉是非姐妹染色单体在同源物之间交换遗传物质的位置。此外，一个CO事件可能会影响沿着染色体附近另一个CO事件的发生，这被称为CO干扰。在过去的几十年里，可视化CO事件和测量CO干扰，大大有助于我们了解减数分裂重组的调控机制。在这里，我们描述的协议，以估计交叉和CO干扰的数目在拟南芥中使用细胞学方法，涉及染色体扩散和免疫染色。具体而言，我们描述了如何使用染色体扩散来估计基于中期I二价体构象的交叉数，并提供了一种修订的基于酸的快速免疫染色试验，该试验允许高通量和定量数字估计终变期相邻干扰敏感CO灶之间的相对距离。这些方法可以很容易地建立或修改，如果需要的话，研究减数分裂重组在其他植物和作物。© 2022 Wiley Periodicals LLC.
Meiosis involves the replication of nuclear chromosomes in a parent cell, followed by two successive nuclear divisions to produce haploid spores, which differentiate into the gametophyte generations that produce the egg and sperm in plants. Meiotic recombination or crossover (CO) is a hallmark of meiosis that allows shuffling of genetic information between homologous chromosomes (homologs), thereby giving rise to genetically diverse progeny cells and, ultimately, individuals in the progeny; this opens vast opportunities for genetic differentiation and hence speciation. Meiotic COs also ensure the formation of bivalents and fidelity of their equal segregation. Therefore, mechanisms that regulate meiotic recombination have been extensively studied in multiple species. Several approaches have been developed to observe or estimate the frequency of CO, in which CO can be visualized and analyzed cytologically by estimating the number of chiasma (plural chiasmata), a position where non‐sister chromatids exchange genetic material between homologs. Furthermore, one CO event might influence the occurrence of another one nearby, along a chromosome; this is known as CO interference. Over the past decades, visualizing CO events and measuring CO interference have contributed greatly to our understanding of the regulatory mechanisms of meiotic recombination. Here, we describe protocols to estimate the number of chiasmata and CO interference in Arabidopsis using cytological methods involving chromosome spreads and immunostaining. Specifically, we describe how chromosome spreads can be used to estimate the number of chiasmata based on the conformations of metaphase I bivalents and provide a revised acid‐based quick immunostaining assay that permits high‐throughput and quantitative digital estimation of the relative distance between adjacent interference‐sensitive CO foci at diakinesis. These methods can be easily established or modified, if necessary, for studying meiotic recombination in other plants and crops. © 2022 Wiley Periodicals LLC.