Isolation and Short-Term Culture of Mouse Spermatocytes for Analysis of Meiosis
Isolation and Short-Term Culture of Mouse Spermatocytes for Analysis of Meiosis
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DOI：
10.1007/978-1-60761-103-5_17
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发表时间：
2009-01-01
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影响因子：
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通讯作者：
Handel, Mary Ann
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文献类型：
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作者：
La Salle, Sophie;Sun, Fengyun;Handel, Mary Ann
体外实验方法有助于理解减数分裂，但这并不容易适用于哺乳动物减数分裂。哺乳动物卵母细胞体外分析的现有方法通常仅限于前期后期的实验分析。雄性生殖细胞的短期培养可用于分析早期减数分裂前期途径以及减数分裂阶段的开始，但没有研究在体外实现可靠的精子发生。在这里，我们描述了一种从小鼠睾丸细胞悬浮液中制备高度富集的粗线期精母细胞的方法，该方法通过在单位重力下通过牛血清白蛋白梯度进行沉降来进行细胞大小分级。我们还提供了精母细胞短期培养和前期到分裂相变的药理学诱导的程序。
Understanding meiosis is facilitated by in vitro experimental approaches, but this has not been easily applicable to mammalian meiocytes. Available methods for in vitro analysis of mammalian oocytes are generally limited to experimental analysis of the late prophase period. Short-term cultures of male germ cells have been useful for analysis of earlier meiotic prophase pathways, as well as onset of the meiotic division phase, but no studies have achieved reliable spermatogenesis in vitro. Here we describe a method for preparing highly enriched pachytene spermatocytes from mouse testicular cell suspensions using cell-size fractionation by sedimentation through a bovine serum albumin gradient at unit gravity. We also provide a procedure for short-term culture of spermatocytes and the pharmacological induction of the prophase-to-division phase transition.