Application of lacZ transgenic mice to cell lineage studies.
Application of lacZ transgenic mice to cell lineage studies.
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lacZ转基因小鼠在细胞谱系研究中的应用。
DOI：
10.1007/978-1-60327-483-8_10
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发表时间：
2008
期刊：
影响因子：
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通讯作者：
Tam,PatrickPL
中科院分区：
文献类型：
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作者：
Watson,CatherineM;Trainor,PaulA;Radziewic,Tania;Pelka,GregoryJ;Zhou,SheilaX;Parameswaran,Maala;Quinlan,GabrielA;Gordon,Monica;Sturm,Karin;Tam,PatrickPL
细胞谱系分析追踪源自祖细胞群体的细胞类型的层级。这些分析的关键是能够可靠地跟踪所有或确定的子集的克隆后代的祖先人口。这需要用可遗传的和细胞自主的标记物标记细胞。最广泛使用的是编码可以直接原位可视化而不损害细胞分化的分子的转基因，例如报告基因、由lacZ编码的β-半乳糖苷酶和由CAT基因编码的氯霉素乙酰转移酶。使用灵敏的组织化学测定法可以很容易地检测到lacZ，使得β-半乳糖苷酶基因具有转录活性的细胞在组织切片或整个标本中产生蓝色染色（1）。一些谱系研究需要同时检测lacZ产物和其他组织特异性蛋白质或转录物。在这一章中，我们讨论了实验策略，其中的lacZ转基因可用于细胞谱系的分析，我们详细分析检测β-半乳糖苷酶的X-gal组织化学或结合mRNA原位杂交，免疫化学和组织化学方法，如碱性磷酸酶染色的酶的免疫定位。
Cell lineage analyses trace the hierarchy of cell types derived from a progenitor population. Critical to these analyses is the ability to track reliably all or defined subsets of the clonal descendants of the progenitor population. This necessitates marking the cells with a heritable and cell autonomous marker. Transgenes encoding molecules that can be visualized directly in situ without compromising cell differentiation, such as the reporter, β-galactosidase encoded by lacZ and chloramphenicol acetyltransferase, encoded by the CAT gene are the most widely used. The lacZ can be readily detected using a sensitive histochemical assay such that cells in which the β-galactosidase gene is transcriptionally active produce a blue stain in tissue sections or in whole mounts (1). Some lineage studies demand the simultaneous detection of the lacZ product and other tissue-specific proteins or transcripts. In this chapter, we discuss the experimental strategies in which the lacZ transgene can be utilized in the analysis of cell lineages, and we detail assays for detecting β-galactosidase by X-gal histochemistry or immunological localization of the enzyme in combination with mRNA in situ hybridization, immunochemistry, and histochemical procedures, such as alkaline phosphatase staining.