Nucleic acid capture, concentration, and purification
An example of a kit includes a flow cell assembly. The flow cell assembly includes a reaction chamber, a temperature controlled flow channel in selective fluid communication with an inlet of the reaction chamber, and a filter positioned in the temperature controlled flow channel. The reaction chamber includes depressions separated by interstitial regions and capture primers attached within each of the depressions. The filter is i) to block concentrated biological sample-polymer complexes generated in the temperature controlled flow channel at a first temperature, and ii) to allow passage of concentrated biological sample and polymer released from the complexes in the temperature controlled flow channel at a second temperature.
1 . A method, comprising:
introducing a sample fluid to a temperature controlled flow channel having a filter positioned therein, the sample fluid including:
an aqueous carrier;
a DNA sample;
methyl cellulose;
a polymer that is chemically inert to DNA hybridization; and
a salt;
heating the temperature controlled flow channel as the sample fluid is introduced such that a temperature of the sample fluid contained therein is increased to at least a gelation temperature of the methyl cellulose, thereby forming DNA-methyl cellulose complexes in the temperature controlled flow channel;
continuing a flow of the sample fluid through the temperature controlled flow channel as the temperature controlled flow channel is heated, thereby concentrating a plurality of the DNA-methyl cellulose complexes at the filter in the temperature controlled flow channel; and
cooling the temperature controlled flow channel such that a temperature of the sample fluid contained therein is decreased to below the gelation temperature of the methyl cellulose, thereby detangling the concentrated DNA-methyl cellulose complexes to release the DNA sample and the methyl cellulose, whereby the DNA sample and the methyl cellulose are able to pass through the filter.
2 . The method as defined in claim 1 , wherein the DNA sample includes cell-free DNA, library DNA, whole genome amplified DNA, or combinations thereof.
3 . The method as defined in claim 1 , further comprising selecting a heating temperature for the temperature controlled flow channel in accordance with a concentration of the salt in the sample fluid.
4 . The method as defined in claim 3 , further comprising increasing the concentration of the salt in the solution, thereby lowering the gelation temperature of the methyl cellulose.
5 . The method as defined in claim 1 , further comprising transporting the DNA sample and the methyl cellulose from the temperature controlled flow channel to a flow cell.
6 . The method as defined in claim 1 , wherein the polymer that is chemically inert to DNA hybridization consists essentially of polyethylene glycol.
7 . The method as defined in claim 6 , wherein the polyethylene glycol has a weight average molecular weight of from 500 grams/mol to 200,000 grams/mol.
8 . The method as defined in claim 1 , wherein the polymer that is chemically inert to DNA hybridization is selected from the group consisting of polyethyleneglycol, polyvinyl pyrrolidone, polyvinyl alcohol, and combinations thereof.
9 . A method, comprising:
introducing a sample fluid to a flow cell, the sample fluid including:
an aqueous carrier;
a DNA sample;
methyl cellulose;
a polymer that is chemically inert to DNA hybridization; and
a salt;
and the flow cell including a reaction chamber having:
depressions separated by interstitial regions; and
capture primers attached within each of the depressions;
initiating seeding and hybridization of at least some of the DNA sample in at least some of the depressions;
heating the flow cell such that a temperature of the sample fluid contained therein is increased to at least a gelation temperature of the methyl cellulose, thereby forming DNA-methyl cellulose complexes with unbound DNA sample;
introducing an additional amount of the sample fluid to the flow cell;
cooling the flow cell such that the temperature of the sample fluid contained therein is decreased to below the gelation temperature of the methyl cellulose, thereby detangling the DNA-methyl cellulose complexes to release the DNA sample and the methyl cellulose; and
initiating seeding and hybridization of at least some of the released DNA sample and the DNA sample from the additional amount of the sample fluid in at least some of the depressions.
10 . The method as defined in claim 9 , further comprising selecting a heating temperature for the flow cell in accordance with a concentration of the salt in the sample fluid.