Emulsion compositions
View Patent ↗An emulsion is useful in allowing a wide variety of gene products to be expressed via eukaryotic in vitro expression. The emulsion comprises a silicone based surfactant, a hydrophobic phase and a hydrophilic phase; wherein the hydrophilic phase comprises a plurality of compartments containing a functional in vitro eukaryotic expression system.
1. A method for increasing the concentration of a nucleic acid molecule comprising the steps of:
(a) forming microcapsules from a water-in-oil emulsion consisting of: a chemically inert silicone-based surfactant, a hydrophobic oil external phase, and a hydrophilic aqueous internal phase comprising the microcapsules, wherein the microcapsules include a nucleic acid molecule, a bead capable of being linked to the nucleic acid molecule, and an aqueous solution comprising components necessary to perform nucleic acid amplification;
(b) amplifying the nucleic acid molecule in the microcapsules to form amplified copies of the nucleic acid molecule, wherein the amplification employs a thermal cycling process, thereby increasing the concentration of the nucleic acid molecule in the microcapsules.
2. The method of claim 1 wherein the amplified copies are captured to the beads in the microcapsules.
3. The method of claim 1 wherein the nucleic acid amplification is performed using a method selected from the group consisting of Qb replicase amplification, ligase chain reaction, self sustained sequence replication, and strand displacement amplification.
4. The method of claim 1 wherein the water-in-oil emulsion comprises a water to oil ratio of 1 to 2.5.
5. The method of claim 1 wherein the nucleic acid molecule is genomic DNA or cDNA.
6. The method of claim 1 wherein the nucleic acid molecule comprises a tag selected from the group consisting of biotin, digoxigenin, and 2,4-dinitrophenyl.
7. The method of claim 1 wherein the bead comprises a coating selected from the group consisting of avidin, streptavidin, anti-digoxigenin antibodies, and anti-2,4-dinitrophenyl antibodies.
8. The method of claim 1 wherein the bead is selected from the group consisting of polystyrene and magnetic beads.
9. The method of claim 1 wherein the chemically inert silicone-based surfactant is provided at a v/v concentration range of 0.5-20% of surfactant in oil.
10. The method of claim 1 wherein the bead with the captured amplification copies is sorted for sequencing.
11. The method of claim 1 wherein a plurality of microcapsules when formed each contains on average one or less than one nucleic acid molecule.
12. The method of claim 1 wherein a plurality of microcapsules when formed each contains on average between 5 and 1000 nucleic acid molecules.