In vitro evolution in microfluidic systems

Patent metadata

Field	Value
Publication number	US-10639598-B2
Application number	US-201815912033-A
Country	US
Kind code	B2
Filing date	Mar 5, 2018
Priority date	Oct 8, 2004
Publication date	May 5, 2020
Grant date	May 5, 2020

Abstract

The invention describes a method for isolating one or more genetic elements encoding a gene product having a desired activity, comprising the steps of: (a) compartmentalising genetic elements into microcapsules; and (b) sorting the genetic elements which express the gene product having the desired activity; wherein at least one step is under microfluidic control. The invention enables the in vitro evolution of nucleic acids and proteins by repeated mutagenesis and iterative applications of the method of the invention.

First claim

What is claimed is:

1. A method for analyzing compounds, the method comprising:
providing a droplet generator comprising a nozzle through which aqueous fluid is flowed under microfluidic control into a reservoir of oil to form a plurality of aqueous microcapsules in the oil, wherein each aqueous microcapsule comprises a genetic element and a cell;
incubating the plurality of aqueous microcapsules to cause the genetic element to interact with a molecule associated with the cell;
conducting an enzymatic reaction involving the molecule associated with the cell within one or more of the aqueous microcapsules, wherein the genetic element or a gene product encoded by the genetic element has an effect on the enzymatic reaction, to produce reaction products corresponding to individual gene products or genetic elements; and
analyzing the individual genetic elements identified based on detection of a selectable change caused by the reaction products in one or more of the aqueous microcapsules.

2. The method of claim 1 , wherein the plurality of aqueous microcapsules are unable to fuse or coalesce due to the presence of a surfactant.

3. The method of claim 1 , wherein the selectable change comprises an optical signal.

4. The method of claim 1 , wherein the genetic element comprises one or more primer sequences for amplification.

5. The method of claim 1 , wherein the genetic element is attached to a bead.

6. The method of claim 1 , wherein the molecule is a reporter protein.

7. The method of claim 1 , wherein the molecule comprises a gene.

8. The method of claim 7 , further comprising expressing the gene to form a gene product.

9. The method of claim 8 , wherein the gene product is a protein.

10. The method of claim 9 , wherein the protein is an antibody.

11. The method of claim 9 , wherein the protein is located within the cell.

12. The method of claim 9 , wherein the protein is located outside of the cell.

13. The method of claim 1 , wherein the molecule is DNA or RNA.

14. The method of claim 9 , wherein the gene product is cDNA.

Assignees

• Res & Innovation Uk
• Harvard College

Inventors

• Griffiths Andrew David
• Weitz David A
• Link Darren Roy
• Ahn Keunho
• Bibette Jerome

Classifications

• B01L3/502746 Primary

  characterised by the means for controlling flow resistance, e.g. flow controllers, baffles or throttle valves · CPC title

• B01L2400/0487

  fluid pressure, pneumatics · CPC title

• B01J2219/00576

  fluorophore · CPC title

• B01J2219/00466

  in a slurry · CPC title

• C12P21/00

  Preparation of peptides or proteins (single cell protein C12N1/00) · CPC title