Nucleic acid extraction using organic solvents to remove inhibitors
Inventors
Chen, Li • STEVENS, JODIE • SCHWAB, KRISTEN • Boone, James • LYERLY, DAVID
Assignees
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Publication Number
US-10196632-B2
Publication Date
2019-02-05
Expiration Date
Abstract
Nucleic acid amplification tests have been widely used in clinical laboratories. Nucleic acid extraction from biological materials is challenging because different unfavorable substances may co-extract and inhibit downstream applications. The present invention relates to a composition of and a method for treating the sample prior, during or post extraction of nucleic acid. More specifically, the claimed invention relates to a composition of and a method for using low concentrations of common organic solvents to remove inhibitors of nucleic acid amplification. The present invention can be used for extracting nucleic acids (DNA/RNA) from bacteria, viruses, parasites, and other biological materials or matrices, including but not limit to, stool samples, body fluids, plants and cultures. The method is rapid, low-cost, and easy to use in a laboratory setting. The nucleic acid extracted in accordance with the invention can be used for nucleic acid amplification reactions.
Core Innovation
The invention provides a method for collecting a biological sample for extraction of nucleic acid from the biological sample. A sample collection device collects the biological sample onto a first portion of the device, and the first portion is inserted through a first end into an interior portion of a sample collection tube. Extraction buffer is added to form a diluted biological sample, where the extraction buffer comprises about 0.5 to about 20 wt. % of an organic solvent selected from acetone, methanol, ethanol, isopropanol, butanol, or a combination thereof.
The diluted biological sample and the first portion of the sample collection device present in the tube are exposed to a temperature of from about 15°C to about 35°C for about 30 minutes or less to extract at least a portion of nucleic acid into the extraction buffer. A filter tip is secured to the first end of the sample collection tube, and the diluted biological sample is filtered such that at least a portion of the nucleic acid and at least a portion of the extraction buffer flow through the filter and are received in a reaction tube.
In a corresponding system, a sample collection device and sample collection tube are provided to receive extraction buffer containing about 0.5 to about 20 wt. % organic solvent. A filter tip allows nucleic acid present in the biological sample to exit the tube by passing through the filter, and a reaction tube is adapted to receive the extraction buffer including nucleic acid for nucleic acid processing. Optional refinements define filter composition and pore characteristics, and tube mechanics that change the interior volume.
Claims Coverage
The excerpt contains two independent claims, a method claim and a system claim. Across these claims, the inventive coverage centers on an extraction buffer with low concentration organic solvent, nucleic-acid extraction at 15–35°C for 30 minutes or less, and a filter tip so nucleic acid and extraction buffer flow into a reaction tube.
Solvent-containing extraction buffer for diluted biological sample
Adding an extraction buffer to the interior portion of the sample collection tube to form a diluted biological sample, the extraction buffer comprising about 0.5 to about 20 wt. % of an organic solvent selected from acetone, methanol, ethanol, isopropanol, butanol, or a combination thereof.
Temperature-limited nucleic-acid extraction
Exposing the diluted biological sample and the first portion of the sample collection device present in the interior portion of the sample collection tube to a temperature of from about 15°C to about 35°C for about 30 minutes or less to extract at least a portion of nucleic acid from the biological sample into the extraction buffer.
Filter-tip flow of nucleic acid and extraction buffer to a reaction tube
Securing a filter tip onto the first end of the sample collection tube, the filter tip comprising a filter, and filtering the diluted biological sample using the filter tip such that at least a portion of the nucleic acid and at least a portion of the extraction buffer flow through the filter and are received in a reaction tube.
System components for tube-based solvent extraction and filtered release
Providing a sample collection device, a sample collection tube defining an interior portion sized to receive the sample collection device and a volume of extraction buffer comprising about 0.5 to about 20 wt. % organic solvent, a filter tip adapted to allow at least a portion of nucleic acid present in the biological sample to exit the interior portion by passing through the filter, and a reaction tube adapted to receive the extraction buffer that includes nucleic acid.
The independent claims collectively cover a method and a system in which nucleic acid is extracted from a biological sample using an extraction buffer containing about 0.5–20 wt. % of specified organic solvents, extraction is performed at 15–35°C for 30 minutes or less, and a filter tip is used so nucleic-acid-containing extraction buffer flows into a reaction tube.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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