Methods and compositions for the extraction and amplification of nucleic acid from a sample
First Claim
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1. A method for extracting a target nucleic acid from a cell-free biological sample, which comprises:
- (a) mixing a cell-free biological sample, a chaotropic salt and a solid support capable of reversibly binding nucleic acid, wherein;
the sample comprises target nucleic acid and non-target nucleic acid,the target nucleic acid and the non-target nucleic acid are both DNA or both RNA,the size of the target nucleic acid is less than the size of the non-target nucleic acid, andthe chaotropic salt is present at a concentration at which the non-target nucleic acid binds to the solid support and substantially no target nucleic acid adsorbs to the solid support, whereby the non-target nucleic acid is bound to the solid support in a first binding solution, wherein the first binding solution does not include alcohol and chloroform;
(b) separating the solid support from the first binding solution, thereby yielding a fraction separated from the solid support;
(c) mixing the fraction separated from the solid support with additional chaotropic salt and an additional solid support capable of reversibly binding nucleic acid, wherein;
the chaotropic salt is present at a concentration at which the target nucleic acid binds to the solid support, whereby the target nucleic acid is bound to the solid support in a second binding solution, wherein the second binding solution does not include alcohol and chloroform and wherein the type of the additional solid support is the same as the type of the solid support in (a);
(d) separating the solid support from the second binding solution; and
(e) eluting the target nucleic acid from the solid support after (d), whereby the target nucleic acid is extracted from the cell-free biological sample.
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Abstract
Provided herein are methods, compositions and kits to extract and relatively enrich by physical separation or amplification short base pair nucleic acid in the presence of a high background of genomic material (e.g., host or maternal nucleic acids).
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Citations
20 Claims
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1. A method for extracting a target nucleic acid from a cell-free biological sample, which comprises:
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(a) mixing a cell-free biological sample, a chaotropic salt and a solid support capable of reversibly binding nucleic acid, wherein; the sample comprises target nucleic acid and non-target nucleic acid, the target nucleic acid and the non-target nucleic acid are both DNA or both RNA, the size of the target nucleic acid is less than the size of the non-target nucleic acid, and the chaotropic salt is present at a concentration at which the non-target nucleic acid binds to the solid support and substantially no target nucleic acid adsorbs to the solid support, whereby the non-target nucleic acid is bound to the solid support in a first binding solution, wherein the first binding solution does not include alcohol and chloroform; (b) separating the solid support from the first binding solution, thereby yielding a fraction separated from the solid support; (c) mixing the fraction separated from the solid support with additional chaotropic salt and an additional solid support capable of reversibly binding nucleic acid, wherein; the chaotropic salt is present at a concentration at which the target nucleic acid binds to the solid support, whereby the target nucleic acid is bound to the solid support in a second binding solution, wherein the second binding solution does not include alcohol and chloroform and wherein the type of the additional solid support is the same as the type of the solid support in (a); (d) separating the solid support from the second binding solution; and (e) eluting the target nucleic acid from the solid support after (d), whereby the target nucleic acid is extracted from the cell-free biological sample. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20)
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Specification