Use of laser capture microdissection for analysis of retinal mRNA/miRNA expression and DNA methylation.
Use of laser capture microdissection for analysis of retinal mRNA/miRNA expression and DNA methylation.
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DOI：
10.1007/978-1-61779-848-1_21
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发表时间：
2012
期刊：
影响因子：
--
通讯作者：
Zack, Donald J
中科院分区：
文献类型：
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作者：
Hackler, Laszlo Jr;Masuda, Tomohiro;Oliver, Verity F;Merbs, Shannath L;Zack, Donald J
激光捕获显微切割（LCM）是从异质组织（例如视网膜）分离感兴趣的特定细胞或细胞层的有用方法。收集的细胞可用于DNA、RNA或蛋白质分析。我们应用LCM技术从视网膜的外核、内核和神经节细胞层分离细胞，用于mRNA和microRNA（miRNA）表达和表观遗传（DNA甲基化）分析。在这里，我们描述了我们所采用的样品制备，基于LCM的视网膜层分离，RNA/DNA提取，RNA质量检查，定量PCR microRNA分析，和亚硫酸氢盐测序DNA甲基化分析的方法。
Laser capture microdissection (LCM) is a useful method to isolate specific cells or cell layers of interest from heterogeneous tissues, such as the retina. The collected cells can be used for DNA, RNA, or protein analysis. We have applied LCM technology to isolate cells from the outer nuclear, inner nuclear, and ganglion cell layers of the retina for mRNA and microRNA (miRNA) expression and epigenetic (DNA methylation) analysis. Here, we describe the methods we have employed for sample preparation, LCM-based isolation of retinal layers, RNA/DNA extraction, RNA quality check, microRNA analysis by quantitative PCR, and DNA methylation analysis by bisulfite sequencing.