US5155018A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Process and kit for isolating and purifying RNA from biological sources
Assignee: Hahnemann University
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Filing: 1991-07-10
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Granted: 1992-10-13
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Expires: 2011-07-10
Abstract
Abstract A process is provided for isolating and purifying biologically active RNA from a biological sources containing RNA, DNA and other cellular materials. The process involves contacting the RNA-containing source with particles comprising siliceous material, such as finely-divided glass, in the presence of a binding solution comprising concentrated, acidified chaotropic salt. Under these conditions, RNA, but not DNA, binds selectively to the siliceous material, and can be separated easily from the other components of the sample. Preferably, the selective binding process is applied to biological cells containing RNA of interest. Intact cells are disrupted by exposing them to a lysing solution comprising, as its main component, concentrated, acidified chaotropic salt. The RNA is then isolated and purified from the lysate using the selective binding process of the invention. The process of the invention may be conveniently provided as a kit, which may include the siliceous material, reagents and instructions for use of the kit to isolate and purify biologically active RNA from biological sources.
Inventors
- 👤 David Gillespie
- 👤 Kevin K. Cuddy
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 19:12
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/10; C12N15/1003; C12N15/1006; B; B01; B01D; B01D15/00; B01D15/08; C12N1/00; C12N1/08; C12Q; C12Q1/00; C12Q1/68; C12Q1/6806; Y; Y10; Y10S; Y10S435/00; Y10S435/803
Claims Analysis
Total Claims: 20
Claim 1
Independent Claim
67 words
A process for isolating biologically active RNA from a biological source containing said RNA, said process comprising: a) contacting said RNA-containing source with particles comprising siliceous material in the presence of an acidified, concentrated chaotropic salt solution, thereby causing RNA present in said source to bind to said particles; b) removing said particle-bound RNA from said source; and c) separating said biologically active RNA from said particles.
Claim 2
Dependent Claim
67 words
A process for isolating biologically active RNA from a biological source containing said RNA, said process comprising: a) contacting said RNA-containing source with particles comprising siliceous material in the presence of an acidified, concentrated chaotropic salt solution, thereby causing RNA present in said source to bind to said particles; b) removing said particle-bound RNA from said source; and c) separating said biologically active RNA from said particles.
Claim 3
Dependent Claim
16 words
The process of claim 1, wherein said biological source is first lysed with a lytic agent.
Claim 4
Dependent Claim
14 words
The process of claim 2, wherein said lytic agent comprises a concentrated chaotropic salt.
Claim 5
Dependent Claim
12 words
The process of claim 3, wherein said chaotropic salt is guanidine thiocyanate.
Claim 6
Dependent Claim
51 words
A process as claimed in claims 1-4, wherein said RNA-containing source is contacted with said particles in the presence of an acidifed, concentrated solution of a chaotropic salt selected from the group consisting of guanidine thiocyanate, guanidine isothiocyanate, guanidine hydrochloride or a mixture of any two or more of said salts.
Claim 7
Dependent Claim
31 words
A process as claimed in claims 1-4, wherein said RNA-containing source is contacted with particles of a siliceous material selected from the group consisting of finely divided glass and diatomaceous earth.
Claim 8
Dependent Claim
23 words
A process as claimed in claims 1-4, wherein said biologically active RNA is isolated from biological cells as the source of said RNA.
Claim 9
Dependent Claim
18 words
A process as claimed in claims 1-4, wherein said acidified chaotropic salt solution is acidifed with acetic acid.
Claim 10
Dependent Claim
17 words
A process as claimed in claims 1-4, wherein said RNA is separated from said particles by elution.
Claim 11
Dependent Claim
37 words
A process as claimed in claim 9, wherein said RNA is eluted from said particles using an eluent comprising a ribonuclease inhibitor, in an amount effective to inhibit residual ribonuclease activity to which said RNA is exposed.
Claim 12
Dependent Claim
31 words
A process as claimed in claim 9, wherein said RNA is eluted from said particles using an eluent comprising deoxyribonuclease, in an amount effective to reduce DNA contamination of said RNA.
Claim 13
Dependent Claim
90 words
A process for obtaining substantially pure, biologically active RNA from a mixture containing RNA and DNA, said process comprising: a) contacting said mixture with particles comprising siliceous material in the presence of an acidified, concentrated chaotropic salt solution, thereby causing RNA present in said source to bind selectively to said particles to the substantial exclusion of said DNA; b) removing said particle-bound RNA from said mixture; c) washing said particle-bound RNA; d) separating said biologically active RNA from said particles; and e) recovering said separated, biologically active, substantially pure RNA.
Claim 14
Dependent Claim
90 words
A process for obtaining substantially pure, biologically active RNA from a mixture containing RNA and DNA, said process comprising: a) contacting said mixture with particles comprising siliceous material in the presence of an acidified, concentrated chaotropic salt solution, thereby causing RNA present in said source to bind selectively to said particles to the substantial exclusion of said DNA; b) removing said particle-bound RNA from said mixture; c) washing said particle-bound RNA; d) separating said biologically active RNA from said particles; and e) recovering said separated, biologically active, substantially pure RNA.
Claim 15
Dependent Claim
39 words
A process as claimed in claim 12, wherein said chaotropic salt solution comprises a chaotropic salt selected from the group consisting of guanidine thiocyanate, guanidine isothiocyanate, guanidine hydrochloride or a mixture of any two or more of said salts.
Claim 16
Dependent Claim
30 words
A process as claimed in claim 12, wherein said mixture is contacted with particles of a siliceous material selected from the group consisting of finely divided glass and diatomaceous earth.
Claim 17
Dependent Claim
61 words
A kit for isolating and purifying, biologically active RNA present in a biological source containing said RNA, comprising: a) a concentrated chaotropic salt solution; b) an acid solution for acidifying said salt solution; c) a suspension of finely divided glass particles; d) an elution solution; and, optionally e) a wash comprising a solution of an inorganic salt in a lower alcohol.
Claim 18
Dependent Claim
61 words
A kit for isolating and purifying, biologically active RNA present in a biological source containing said RNA, comprising: a) a concentrated chaotropic salt solution; b) an acid solution for acidifying said salt solution; c) a suspension of finely divided glass particles; d) an elution solution; and, optionally e) a wash comprising a solution of an inorganic salt in a lower alcohol.
Claim 19
Dependent Claim
22 words
A kit according to claim 15, which further comprises a buffer solution comprising a thiol reducing agent and, optionally, a compatible detergent.
Claim 20
Dependent Claim
17 words
A kit according to claim 15, wherein the components of said kit are provided in single-use aliquots.
References Cited (Backward Citations)
No citation records indexed.
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1995-10-13 | 1996-04-13 | 1996-10-13 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1999-10-13 | 2000-04-13 | 2000-10-13 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2003-10-13 | 2004-04-13 | 2004-10-13 | $7700.00 | Window Closed / Lapsed |