RNAsnap™: a rapid, quantitative and inexpensive, method for isolating total RNA from bacteria.
RNAsnap™: a rapid, quantitative and inexpensive, method for isolating total RNA from bacteria.
复制标题
DOI：
10.1093/nar/gks680
复制
发表时间：
2012-11-01
影响因子：
14.9
通讯作者：
Kushner SR
中科院分区：
文献类型：
--
作者：
Stead MB;Agrawal A;Bowden KE;Nasir R;Mohanty BK;Meagher RB;Kushner SR
RNAsnap™是一种简单而新颖的方法，它可以定量地回收所有细胞内RNA（>99%），比目前可用的任何RNA分离方法更快（<15分钟）且更便宜（0.03美分/样品）。事实上，没有一种细菌RNA分离方法，包括商业试剂盒，能够以相同的效率有效回收所有种类的细胞内RNA（76-5700 nt），这可能导致涉及微阵列或RNAseq分析的全基因组研究中的偏倚结果。RNAsnap™程序从108个大肠杆菌细胞中产生约60 μg RNA，无需进一步纯化即可直接用于北方分析。基于大小范围从76至5700 nt的特定转录物的比较分析，RNAsnap™方法提供了各种细胞内RNA的相对量的最准确测量。此外，RNAsnap™ RNA成功地用于酶促反应，如RNA连接、逆转录、引物延伸和逆转录酶-聚合酶链式反应，随后进行乙酸钠/乙醇沉淀。RNAsnap™方法可用于从多种革兰氏阴性和革兰氏阳性细菌以及酵母中分离RNA。
RNAsnap™ is a simple and novel method that recovers all intracellular RNA quantitatively (>99%), faster (<15 min) and less expensively (∼3 cents/sample) than any of the currently available RNA isolation methods. In fact, none of the bacterial RNA isolation methods, including the commercial kits, are effective in recovering all species of intracellular RNAs (76–5700 nt) with equal efficiency, which can lead to biased results in genome-wide studies involving microarray or RNAseq analysis. The RNAsnap™ procedure yields ∼60 µg of RNA from 108 Escherichia coli cells that can be used directly for northern analysis without any further purification. Based on a comparative analysis of specific transcripts ranging in size from 76 to 5700 nt, the RNAsnap™ method provided the most accurate measure of the relative amounts of the various intracellular RNAs. Furthermore, the RNAsnap™ RNA was successfully used in enzymatic reactions such as RNA ligation, reverse transcription, primer extension and reverse transcriptase–polymerase chain reaction, following sodium acetate/ethanol precipitation. The RNAsnap™ method can be used to isolate RNA from a wide range of Gram-negative and Gram-positive bacteria as well as yeast.
登录
查看更多内容
DOI：
10.1007/bf00330395
发表时间：
1986-01-01
期刊：
MOLECULAR AND GENERAL GENETICS
影响因子：
--
作者：
LINCHAO, S;BREMER, H
通讯作者：
BREMER, H
影响因子：
2.9
作者：
GLISIN, V;CRKVENJAKOV, R;BYUS, C
通讯作者：
BYUS, C
影响因子：
14.9
作者：
Mohanty, Bijoy K.;Kushner, Sidney R.
通讯作者：
Kushner, Sidney R.
影响因子：
14.9
作者：
Stead, Mark B.;Marshburn, Sarah;Kushner, Sidney R.
通讯作者：
Kushner, Sidney R.
影响因子：
2.2
作者：
Jahn, Courtney E.;Charkowski, Amy O.;Willis, David K.
通讯作者：
Willis, David K.