Method for processing polynucleotide-containing samples
DCFirst Claim
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1. A method for processing polynucleotide-containing sample, the method comprising:
- retaining polynucleotides from a sample on a plurality of binding particles in a process chamber under a first set of conditions, wherein the retaining step comprises binding the polynucleotides to the surfaces of the plurality of binding particles comprising a poly-cationic substance, wherein the sample has a volume from 0.5 microliters to 3 milliliters;
wherein the first set of conditions includes a first pH of about 8.5 or less and a first temperature, wherein the first temperature is about 50°
C.;
releasing the polynucleotide from the plurality of binding particles under a second set of conditions; and
wherein the second set of conditions includes increasing the pH to a second pH by addition of a hydroxide solution and increasing the temperature to a second temperature, wherein the second temperature is between about 80°
C. and about 100°
C.
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Abstract
Methods and systems for processing polynucleotides (e.g., DNA) are disclosed. A processing region includes one or more surfaces (e.g., particle surfaces) modified with ligands that retain polynucleotides under a first set of conditions (e.g., temperature and pH) and release the polynucleotides under a second set of conditions (e.g., higher temperature and/or more basic pH). The processing region can be used to, for example, concentrate polynucleotides of a sample and/or separate inhibitors of amplification reactions from the polynucleotides. Microfluidic devices with a processing region are disclosed.
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Citations
20 Claims
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1. A method for processing polynucleotide-containing sample, the method comprising:
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retaining polynucleotides from a sample on a plurality of binding particles in a process chamber under a first set of conditions, wherein the retaining step comprises binding the polynucleotides to the surfaces of the plurality of binding particles comprising a poly-cationic substance, wherein the sample has a volume from 0.5 microliters to 3 milliliters; wherein the first set of conditions includes a first pH of about 8.5 or less and a first temperature, wherein the first temperature is about 50°
C.;releasing the polynucleotide from the plurality of binding particles under a second set of conditions; and wherein the second set of conditions includes increasing the pH to a second pH by addition of a hydroxide solution and increasing the temperature to a second temperature, wherein the second temperature is between about 80°
C. and about 100°
C. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13)
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14. A method for processing polynucleotide-containing samples, the method comprising:
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retaining one or more polynucleotides from a sample on a plurality of binding particles under a first set of conditions, wherein a surface of one or more binding particles is modified with a poly-cationic material, wherein the sample has a volume from 0.5 microliters to 3 milliliters; wherein the first set of conditions includes a first pH of 8.5 or less and a first temperature of about 50°
C.;releasing the polynucleotide from the plurality of binding particles under a second set of conditions; and wherein the second set of conditions includes increasing the pH by at least three units by addition of a hydroxide solution and increasing the temperature by at least about 40°
C. to a second temperature of at least about 90°
C. - View Dependent Claims (15, 16, 17)
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18. A method for processing a polynucleotide-containing sample, the method comprising:
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contacting the sample with a plurality of binding particles, the binding particles retaining one or more polynucleotides thereon at a first pH and a first temperature, wherein the sample has a volume from 0.5 microliters to 3 milliliters, wherein the first temperature is about 50°
C., wherein a surface of one or more binding particles is modified with a poly-cationic polyimide or polyethyleneimine (PEI); andcontacting the binding particles with a basic hydroxide solution at a second pH and a second temperature, the second temperature greater than the first temperature, thereby releasing the polynucleotides from the plurality of binding particle, wherein the second temperature is between about 80°
C. and about 100°
C. - View Dependent Claims (19, 20)
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Specification