Emulsion compositions

• US 20050064460A1
• Filed: 06/11/2004
• Published: 03/24/2005
• Est. Priority Date: 11/16/2001
• Status: Active Grant

• Current Assignee: UK Research and Innovation
• Original Assignee: Medical Research Council (Government of United Kingdom)
• Inventors: Holliger, Phillip, Ghadessy, Farid
• Granted Patent: US 7,622,280 B2
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: A61K 47/24 containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
• CPC Class Codes: A61K 47/26 Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
• CPC Class Codes: A61K 47/44 Oils, fats or waxes according to two or more groups of A61K47/02-A61K47/42; Natural or modified natural oils, fats or waxes, e.g. castor oil, polyethoxylated castor oil, montan wax, lignite, shellac, rosin, beeswax or lanolin synthetic glycerides, e.g. medium-chain triglycerides, A61K47/14
• CPC Class Codes: A61K 9/107 Emulsions ; Emulsion preconcentrates; Micelles composition of emulsions A61K47/00
• CPC Class Codes: A61K 9/1676 having a drug-free core with discrete complete coating layer containing drug adsorbates of liquid drug formulations on inert powders without simultaneous granulation step A61K9/141; with further drug-free outer coating A61K9/5078; drug conjugated to non-active particles A61K47/6921
• CPC Class Codes: C12N 15/1075 by coupling phenotype to genotype, not provided for in other groups of this subclass
• CPC Class Codes: C12Q 1/6844 Nucleic acid amplification reactions
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer

Abstract

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.

Claims

1. (canceled).

2. A method for increasing concentration of a nucleic acid molecule comprising the steps of:

(a) forming microcapsules from a water-in-oil emulsion, 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 and a chemically inert silicone-based surfactant;

(b) amplifying the nucleic acid molecule in the microcapsules to form amplified copies of the nucleic acid molecule,

thereby increasing the concentration of the nucleic acid molecule in the microcapsules.

3. The method of claim 2 wherein the amplified copies are captured to the beads in the microcapsules.

4. The method of claim 2 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.

5. The method of claim 2 wherein the nucleic acid amplification is performed using polymerase chain reaction.

6. The method of claim 2 wherein the water-in-oil emulsion comprises a water to oil ratio of 1 to 2.5.

7. The method of claim 2 wherein the surfactant is a silicone copolymer.

8. The method of claim 2 wherein the surfactant is a polysiloxane-polycetyl-polyethylene glycol copolymer.

9. The method of claim 2 wherein the surfactant is Abil EM90.

10. The method of claim 2 wherein the emulsion is thermostable.

11. The method of claim 2 wherein the nucleic acid molecule is genomic DNA or cDNA.

12. The method of claim 2 wherein the nucleic acid molecule comprises a tag selected from the group consisting of biotin, digoxigenin, and 2,4-dinitrophenyl.

13. The method of claim 2 wherein the bead comprises a coating selected from the group consisting of avidin, streptavidin, anti-digoxigenin antibodies, and anti-2,4-dinitrophenyl antibodies.

14. The method of claim 2 wherein the bead is selected from the group consisting of polystyrene and magnetic beads.

15. The method of claim 2 wherein the surfactant is provided at a v/v concentration in the oil phase of the emulsion of 0.5 to 20%.

16. The method of claim 2 wherein the bead with the captured amplification copies is sorted for sequencing.

17. The method of claim 2 wherein a plurality of microcapsules when formed each contains on average one or less than one nucleic acid molecule.

18. The method of claim 2 wherein a plurality of microcapsules when formed each contains on average between 5 and 1000 nucleic acid molecules.