Abstract
The study of messenger RNA in mammalian cells by Northern analysis requires the extraction of intact RNA in pure form. Although a number of reliable techniques have been developed for the purpose, most are fairly complex, involving steps such as ultracentrifugation and multiple extractions with large volumes of phenol and chloroform. When the number of cell samples to be analyzed is large, these techniques can be unwieldy. I now describe an RNA purification procedure which is simple enough to allow handling of a large number of cultured cell samples. It uses safe and inexpensive reagents and produces a high yield of pure total cell RNA, essentially free of DNA and ribonuclease, suitable for Northern analysis. The procedure also allows extraction of intact RNA from human granulocytes, cells which are rich in ribonuclease and contain very low amounts of RNA.
MeSH Terms
Animals
Cells, Cultured
Electrophoresis, Agar Gel/methods
Granulocytes/analysis
Humans
Indicators and Reagents
Mice
Nucleic Acid Hybridization
RNA/blood,genetics,isolation & purification
RNA, Messenger/genetics
Chemicals
Indicators and Reagents
RNA, Messenger
RNA
Authors & Affiliations
1 authors, click to expand affiliations / ORCID
Birnboim H C
Ottawa Regional Cancer Centre, University of Ottawa, Ontario, Canada.
References (12)
12 references, click to expand
-
Purification of mouse immunoglobulin heavy-chain messenger RNAs from total myeloma tumor RNA.
Eur J Biochem. 1980 Jun;107(2):303-14
PMID: 6772444
-
Quantitative studies of the growth of mouse embryo cells in culture and their development into established lines.
J Cell Biol. 1963 May;17:299-313
PMID: 13985244
-
Molecular structure and evolutionary origin of human cardiac muscle actin gene.
Proc Natl Acad Sci U S A. 1982 Oct;79(19):5901-5
PMID: 6310553
-
Fibronectin gene expression in polymorphonuclear leukocytes. Accumulation of mRNA in inflammatory cells.
J Biol Chem. 1987 Feb 15;262(5):2111-5
PMID: 3818588
-
Isolation and fractionation of total nucleic acids from tissues and cells.
J Biochem Biophys Methods. 1986 Jan;12(1-2):29-36
PMID: 3944418
-
Purification of functional RNA from human granulocytes.
J Lab Clin Med. 1985 Jan;105(1):94-8
PMID: 2578534
-
A method for isolation of intact, translationally active ribonucleic acid.
DNA. 1983;2(4):329-35
PMID: 6198133
-
Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes.
Proc Natl Acad Sci U S A. 1977 Dec;74(12):5350-4
PMID: 414220
-
Co-existence of vinculin and a vinculin-like protein of higher molecular weight in smooth muscle.
J Biol Chem. 1982 Sep 25;257(18):11024-31
PMID: 6809764
-
Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction.
Anal Biochem. 1987 Apr;162(1):156-9
PMID: 2440339
-
DNA strand breakage in human leukocytes exposed to a tumor promoter, phorbol myristate acetate.
Science. 1982 Mar 5;215(4537):1247-9
PMID: 6276978
-
Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease.
Biochemistry. 1979 Nov 27;18(24):5294-9
PMID: 518835