Purifying Hybridized RNA Polynucleotides Using A Washing Buffer With No Ethanol To Remove Unbound Oligonucleotides

A method of purifying hybridized RNA polynucleotides, comprising:

• adding a hybridization solution to a binding buffer and beads, wherein the hybridization solution includes the hybridized RNA polynucleotides that are hybridized to DNA oligonucleotides;
• aggregating the beads using a magnetic field;
• removing the binding buffer and the hybridization solution from the aggregated beads;
• adding a washing buffer to the beads after the binding buffer and the hybridization solution have been removed, wherein the washing buffer includes less than 5% ethanol;
• aggregating the beads to which the washing buffer has been added;
• removing the washing buffer from the aggregated beads;
• adding an elution buffer to the beads after the washing buffer has been removed;
• aggregating the beads to which the elution buffer has been added;
• and
• removing the elution buffer from the aggregated beads, wherein the elution buffer contains purified hybridized RNA polynucleotides from which substantially all DNA oligonucleotides that are not hybridized to RNA polynucleotides have been removed.

A method of purifying hybridized RNA polynucleotides, comprising:

• adding a hybridization solution to a binding buffer and beads, wherein the hybridization solution includes the hybridized RNA polynucleotides that are hybridized to DNA oligonucleotides, and wherein a first bonding site with a first nucleotide sequence on the RNA polynucleotides is hybridized to a second bonding site with a second nucleotide sequence on the DNA oligonucleotides;
• aggregating the beads using a magnetic field;
• removing the binding buffer and the hybridization solution from the aggregated beads;
• adding a washing buffer to the beads after the binding buffer and the hybridization solution have been removed, wherein the washing buffer has a temperature and a stringency, and wherein the stringency of the washing buffer is adjusted to provide a melting temperature between the first bonding site with the first nucleotide sequence and the second bonding site with the second nucleotide sequence of 5 to 10 degrees Celsius above the temperature of the washing buffer;
• aggregating the beads to which the washing buffer has been added;
• removing the washing buffer from the aggregated beads;
• adding an elution buffer to the beads after the washing buffer has been removed;
• aggregating the beads to which the elution buffer has been added;
• and
• removing the elution buffer from the aggregated beads, wherein the elution buffer contains purified hybridized RNA polynucleotides from which substantially all DNA oligonucleotides that are not hybridized to RNA polynucleotides have been removed.