高分辨率细胞谱系追踪揭示了水蛭的发育变异性。
High resolution cell lineage tracing reveals developmental variability in leech.
机构信息
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3200, USA.
出版信息
Dev Dyn. 2009 Dec;238(12):3139-51. doi: 10.1002/dvdy.22158.
Abstract
Knowing the normal patterns of embryonic cell proliferation, migration, and differentiation is a cornerstone for understanding development. Yet for most species, the precision with which embryonic cell lineages can be determined is limited by technical considerations (the large numbers of cells, extended developmental times, opacity of the embryos), and these are exacerbated by the inherent variability of the lineages themselves. Here, we present an improved method of cell lineage tracing in the leech Helobdella, driving the expression of a nuclearly localized histone H2B:GFP (green fluorescent protein) fusion protein in selected lineages by microinjection of a plasmid vector. This construct generates a long lasting and minimally mosaic signal with single cell resolution, and does not disrupt the development of most lineages tested. We have validated this technique by elucidating details of cell lineages contributing to segmental and prostomial tissues that could not be observed with standard dextran lineage tracers.
摘要
了解胚胎细胞增殖、迁移和分化的正常模式是理解发育的基石。然而，对于大多数物种来说，确定胚胎细胞谱系的精确性受到技术考虑因素的限制（细胞数量庞大、发育时间延长、胚胎不透明），而这些因素又因谱系本身的固有可变性而加剧。在这里，我们提出了一种改进的秀丽隐杆线虫（Helobdella）细胞谱系追踪方法，通过微注射质粒载体在选定的谱系中驱动核定位组蛋白 H2B：GFP（绿色荧光蛋白）融合蛋白的表达。这种构建体产生具有单细胞分辨率的持久且最小程度镶嵌的信号，并且不会破坏大多数测试谱系的发育。我们通过阐明对节段和前体组织有贡献的细胞谱系的细节来验证这种技术，这些细节是用标准葡聚糖谱系示踪剂无法观察到的。
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