US 5,393,672 AGranted

Non toxic compositions and methods useful for the extraction of nucleic acids

AssigneeMicroProbe, Inc.

InventorsJeffrey V. Ness, B. Melina Cimler, Rich B. Meyer, Jr., Nicolaas M. J. Vermeulen

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Abstract

This invention relates to safe and effective methods for the extraction of nucleic acids. In particular, methods are described for isolating nucleic acid from a sample containing a biological mixture of nucleic acids and other biological compounds wherein the sample is combined with an extraction solution containing at least one organic compound such as benzyl alcohol or a benzyl alcohol derivative to form an aqueous and non-aqueous phase. The nucleic acid is isolated from the aqueous phase. Preferably, the resulting combined solution also contains bentonite, as defined below. Typically, the sample will first be combined with a lysing agent before extraction. The lysing agents preferred are chaotropic salts such as guanidinium hydrochloride and guanidinium isothiocyanate.

Claims25

Dependent claims

An extraction composition for the isolation of released nucleic acids comprising:

at least one organic compound which is a liquid at room temperature and which is selected from the group of compounds having the formula C.sub.6 H.sub.5 CH.sub.2 OH or:
##STR2## where:
R.sup.1 is --CH.sub.2 OH, R.sup.2 is a member selected from the group consisting of --H, --F, --C, --Br, --I, --OH, --S(CH.sub.2).sub.r CH.sub.3 in which r is 0 to 3, --O(CH.sub.2).sub.q CH.sub.3 in which q is 0 to 6, --COOR.sup.6 in which R.sup.6 is --(CH.sub.2).sub.s CH.sub.3 in which s is 0 to 3, --C.sub.6 H.sub.5, --CH.sub.2 C.sub.6 H.sub.5 , --OC.sub.6 H.sub.5, --OCH.sub.2 C.sub.6 H.sub.5, --CH.sub.2 OH, --CF.sub.3, and --(CH.sub.2).sub.t CH.sub.3 in which t is 0 to 6 and R.sup.3 and R.sup.4 are both --H;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2 and R.sup.3 are the same member selected from the group consisting of --H, --F, --Cl, --Br, --I, --OH, --(CH.sub.2).sub.x CH.sub.3 in which x is 0 to 3, --O(CH.sub.2).sub.v CH.sub.3 in which v is 0 to 3 and R.sup.4 is --H;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2, R.sup.3 and R.sup.4 are the same member selected from the group consisting of --H, --F, --Cl, --Br, --I, --(CH.sub.2).sub.y CH.sub.3 in which y is 0 to 3, --O(--CH.sub.2).sub.w CH.sub.3 in which w is 0 to 3;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2 is --C.sub.6 H.sub.5, --OCH.sub.2 C.sub.6 H.sub.5, or --CH.sub.2 C.sub.6 H.sub.5, R.sup.3 is --CH.sub.3, --OH, --Cl, --Br, --F, --I, --H, --OCH.sub.3 or --OCH.sub.2 CH.sub.3 and R.sup.4 is --H;
and
a clay or a silicate or an admixture of said clay and said silicate;
wherein said clay or said silicate or said admixture is 0.1% to about 5% by weight of said composition.

9 more elements

An extraction composition for the isolation of released nucleic acids comprising:

a clay or a silicate or an admixture of said clay and said silicate;
and
at least one organic compound which is selected from the group of compounds having the formula C.sub.6 H.sub.5 CH.sub.2 OH or:
##STR3## where:
R.sup.1 is --CH.sub.2 OH;
R.sup.2 is a member selected from the group consisting of --H, --F, --Cl, --Br, --I, --OH, --S(CH.sub.2).sub.r CH.sub.3 in which r is 0 to 3, --O(CH.sub.2).sub.q CH.sub.3 in which q is 0 to 6, --COOR.sup.6 in which R.sup.6 is --(CH.sub.2).sub.s CH.sub.3 in which s is 0 to 3, --C.sub.6 H.sub.5, --CH.sub.2 C.sub.6 H.sub.5, --OC.sub.6 H.sub.5, --OCH.sub.2 C.sub.6 H.sub.5, --CH.sub.2 OH, --CF.sub.3, and --(CH.sub.2).sub.t CH.sub.3 in which t is 0 to 6 and R.sup.3 and R.sup.4 are both --H;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2 and R.sup.3 are the same member selected from the group consisting of --H, --F, --Cl, --Br, --I, --OH, --(CH.sub.2).sub.x CH.sub.3 in which x is 0 to 3, --O(CH.sub.2).sub.v CH.sub.3 in which v is 0 to 3 and R.sup.4 is --H;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2, R.sup.3 and R.sup.4 are the same member selected from the group consisting of --H, --F, --Cl, --Br, --I, --(CH.sub.2).sub.y CH.sub.3 in which y is 0 to 3, --O(CH.sub.2).sub.w CH.sub.3 in which w is 0 to 3;
or
R.sup.1 is --CH.sub.2 OH, R.sup.2 is --C.sub.6 H.sub.5, --OCH.sub.2 C.sub.6 H.sub.5, or --CH.sub.2 C.sub.6 H.sub.5, R.sup.3 is --CH.sub.3, --OH, --Cl, --Br, --F, --I, --H, --OCH.sub.3 or --OCH.sub.2 CH.sub.3 and R.sup.4 is --H;
and
wherein the organic compound selected is one that is a solid at room temperature which is dissolved in an organic solvent.

11 more elements

Dependencies are inferred from the claim text.

Facts

Application: 08/156,519Filed 23 Nov 1993 · continuation
Priority: 13 Jul 1990
Granted: 28 Feb 1995
Term ends: Not estimatedThe source records don’t support an estimate.

Classification8 codes

C12Q 1/6806
Involving nucleic acids: preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
C12N 15/1003
Processes for the isolation, preparation or purification of DNA or RNA: extracting or separating nucleic acids from biological samples, e.g. pure…
Y10T 436/143333
Hetero-O [e.g., ascorbic acid, etc.]: saccharide [e.g., DNA, etc.]
Y10T 436/25125
Including sample preparation: digestion or removing interfering materials

Family

Serial552,745Parent of this continuation-in-part · filed 13 Jul 1990
Serial649,389Parent of this continuation · filed 1 Feb 1991
Serial900,379Parent of this continuation · filed 17 Jun 1992
US 5,130,423 AParent of this continuation
US 5,393,672 AThis patent

From the US filing history. Foreign family members aren’t listed.