Immobilization of message RNA directly from cells onto filter ma

Details Immobilization of message RNA directly from cells onto filter ma Immobilization of message RNA directly from cells onto filter ma

1982-05-17

1984-11-20

Naff, David M.

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving nucleic acid

435 92, 4351722, 435174, 435179, C12Q 168, C12P 1934, C12N 1500, C12N 1112

Patent

active

044839200

ABSTRACT:
Message RNA is immobilized directly from cells onto filter material. Immobilization is carried out by solubilizing cellular components with a chaotropic salt, passing the resultant solubilized cellular components through a filter which selectively binds message RNA and baking the filter containing bound message RNA. The chaotropic salt is preferably sodium iodide, potassium iodide or sodium perchlorate. Prior to solubilizing, the cells may be washed and lysed. The bound message RNA can be hybridized to a labeled probe and the amount of message RNA measured. Prior to baking, the filter containing bound RNA may be incubated in a solution which acelylates basic protein and other molecules which might interfere with molecular hybridization.

REFERENCES:
patent: 4302204 (1981-11-01), Wahl et al.
patent: 4358535 (1982-11-01), Falkow et al.
Gilham, P. T., The Synthesis of Celluloses Containing Covalently Bound Nucleotides, Polynucleotides and Nucleic Acids, Biochemistry, vol. 7, No. 8, 1968 (pp. 2809-2813).
Thomas, P. S., Hybridization of Denatured RNA and Small DNA Fragments Transferred to Nitrocellulose, Proc. Nat'l. Acad. Sci. U.S.A., vol. 77, No. 9, 1980 (pp. 5201-5205).
Benton, et al., Screening .lambda.gt Recombinant Clones by Hybridization to Single Plaques in situ, Science, vol. 196, 1977 (pp. 180-182).
Thang, et al., Observations on the Activity of Enzymes after Filtration on (and through) a Nitrocellulose Membrane, Biochem. and Biophy. Res. Comm., vol. 31, No. 1, 1968 (pp. 1-8).
Poonian, et al., Covalent Attachment of Nucleic Acids to Agarose for Affinity Chromatography, Biochemistry, vol. 10, No. 3, 1971 (pp. 424-427).
Journal of the American Chemical Society, Apr. 27, 1962, pp. 1329-1338.

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