Cytoplasmic RNA extraction from fresh and frozen mammalian tissues
Cytoplasmic RNA extraction from fresh and frozen mammalian tissues
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DOI：
10.2144/02332st01
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发表时间：
2002-08-01
期刊：
影响因子：
2.7
通讯作者：
Brownstein, MJ
中科院分区：
文献类型：
--
作者：
Carninci, P;Nakamura, M;Brownstein, MJ
表达序列标签和全长 cDNA 集合的质量受到来自常规总 RNA 制剂中存在的未剪接或部分剪接 RNA 的“垃圾”克隆的存在的不利影响。人们可以通过使用完整的细胞质 RNA 创建 cDNA 文库来克服这一问题，但文献中描述的 RNA 制备方法仅适用于提取培养细胞。细胞系并不像人们希望的那样多样化，为了克隆全套人类和模型生物全长 cDNA，必须从组织样本中制备文库。因此，我们开发了一种稳健且廉价的方法，可以从新鲜和冷冻中提取完整的细胞质 RNA。哺乳动物组织。使用这种新方法用 RNA 制备的小鼠全长、带帽捕获的 cDNA 文库具有优异的特性。
The quality of collections of expressed sequence tags and full-length cDNAs is adversely affected by the presence of "junk" clones derived from unspliced or partially spliced RNAs present in conventional total RNA preparations. One can overcome this problem by using intact cytoplasmic RNA to create cDNA libraries, but the methods in the literature that describe the preparation of RNA only work well for extracting cultured cells. Cell lines are not as diverse as one would like, and to clone comprehensive sets of human and model organism full-length cDNAs, libraries have to be prepared from tissue samples. Thus, we have developed a robust and inexpensive method that allows intact cytoplasmic RNA to be extracted from both fresh and frozen. mammalian tissues. A mouse full-length, cap-trapped cDNA library prepared with RNA using this new procedure had excellent characteristics.