Automated cell processing methods, modules, instruments, and systems

• US 10,519,437 B1
• Filed: 09/14/2019
• Issued: 12/31/2019
• Est. Priority Date: 06/30/2017
• Status: Active Grant

• Current Assignee: Inscripta, Inc.
• Original Assignee: Inscripta, Inc.
• Inventors: Masquelier, Don, Belgrader, Phillip, Bernate, Jorge, Gill, Ryan, Ness, Kevin
• Primary Examiner(s): Bowers, Nathan A
• Application Number:
  US16/571,091
• Time in Patent Office: 108 Days
• CPC Class Codes: C12M 23/44 Multiple separable units; Modules
• CPC Class Codes: C12M 33/14 with filters, sieves or membranes
• CPC Class Codes: C12M 35/00 Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion machines for extracting juice from animal or plant tissue by electroplasmolysis A23N1/006, processes employing electric or wave energy B01J19/08; treatment of microorganisms or enzymes with electrical or wave energy C12N13/00; methods for cell fusion C12N15/02; introduction of foreign genetic material C12N15/87
• CPC Class Codes: C12M 37/04 Seals
• CPC Class Codes: C12M 41/36 of biomass, e.g. colony counters or by turbidity measurements electrooptical investigation of individual particles G01N15/14, flow cytometers G01N15/1404
• CPC Class Codes: C12M 41/48 Automatic or computerized control automatic analysis G01N35/00
• CPC Class Codes: C12M 43/00 Combinations of bioreactors or fermenters with other apparatus
• CPC Class Codes: C12N 15/102 Mutagenizing nucleic acids
• CPC Class Codes: C12N 15/1082 Preparation or screening gene libraries by chromosomal integration of polynucleotide sequences, HR-, site-specific-recombination, transposons, viral vectors
• CPC Class Codes: C12N 15/11 DNA or RNA fragments; Modified forms thereof DNA or RNA not used in recombinant technology, C07H21/00; Non-coding nucleic acids having a biological activity
• CPC Class Codes: C12N 2310/20 involving clustered regularly interspaced short palindromic repeats [CRISPRs]
• CPC Class Codes: C12N 2800/80 Vectors containing sites for inducing double-stranded breaks, e.g. meganuclease restriction sites
• CPC Class Codes: C12N 9/22 Ribonucleases RNAses, DNAses catalytic nucleic acids C12N15/113
• CPC Class Codes: C12Q 1/6865 Promoter-based amplification, e.g. nucleic acid sequence amplification [NASBA], self-sustained sequence replication [3SR] or transcription-based amplification system [TAS]
• CPC Class Codes: C40B 40/02 Libraries contained in or displayed by microorganisms, e.g. bacteria or animal cells; Libraries contained in or displayed by vectors, e.g. plasmids; Libraries containing only microorganisms or vectors

Abstract

In an illustrative embodiment, automated multi-module cell editing instruments are provided to automate multiple edits into nucleic acid sequences inside one or more cells.

Claims

1. An automated stand-alone multi-module cell editing instrument for performing nuclease editing comprising:

a receptacle configured to receive cells and/or nucleic acids;

a growth module for growing the cells for transformation;

a cell concentration module to concentrate the cells and render the cells electrocompetent;

a transformation module configured to introduce nucleic acids into the cells;

a selection module;

a nuclease-directed editing module configured to allow the introduced nucleic acids to edit nucleic acids in the cells;

a processor configured to operate the automated multi-module cell editing instrument based on user input and/or selection of a pre-programmed script; and

an automated liquid handling system to move liquids from the receptacle to the growth module, from the growth module to the cell concentration module, from the cell concentration module to the transformation module, from the transformation module to the selection module, and from the selection module to the nuclease-directed editing module, without user intervention.

2. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the nuclease-directed editing module is also a selection module.

3. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the selection module is separate from the nuclease-directed editing module.

4. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the selection module comprises an antibiotic.

5. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the nuclease editing system employs an RNA-guided nuclease.

6. The automated stand-alone multi-module cell editing instrument of claim 5, wherein the RNA-guided nuclease is MAD7.

7. The automated stand-alone multi-module cell editing instrument of claim 6, wherein the nuclease editing system is an inducible system.

8. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the nuclease editing system employs a zinc-finger nuclease.

9. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the cell concentration module comprises a filter.

10. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the cell concentration module comprises a centrifuge.

11. An automated stand-alone multi-module cell editing instrument for performing nuclease editing comprising:

a receptacle configured to receive cells/or nucleic acids;

a transformation module comprising an electroporator to introduce the nucleic acids into the cells;

a selection module;

a nuclease-directed editing module configured to select for transformed cells and to allow the nucleic acids to edit nucleic acids in the cells to produce edited cells;

a processor configured to operate the automated multi-module cell editing instrument based on user input and/or selection of a script; and

an automated liquid handling system to move liquids from the receptacle to the transformation module, from the transformation module to the selection module, and from the selection module to the nuclease-directed editing module, all without user intervention.

12. The automated stand-alone multi-module cell editing instrument of claim 11, wherein the nuclease editing system employs an RNA-guided nuclease.

13. The automated stand-alone multi-module cell editing instrument of claim 12, wherein the RNA-guided nuclease is MAD7.

14. The automated stand-alone multi-module cell editing instrument of claim 13, wherein the nuclease editing system is inducible.

15. The automated stand-alone multi-module cell editing instrument of claim 11, wherein the nuclease editing system employs a zinc finger nuclease.

16. The automated stand-alone multi-module cell editing instrument of claim 11, wherein the selection module is also the nuclease-directed editing module.

17. The automated stand-alone multi-module cell editing instrument of claim 11, wherein the selection module is separate from the nuclease-directed editing module.

18. The automated stand-alone multi-module cell editing instrument of claim 11, further comprising a cell concentration module.

19. The automated stand-alone multi-module cell editing instrument of claim 18, wherein the cell concentration module comprises a centrifuge.

20. The automated stand-alone multi-module cell editing instrument of claim 18, wherein the cell concentration module comprises a filter.