Automated cell processing methods, modules, instruments, and systems

• US 10,329,559 B1
• Filed: 02/07/2019
• Issued: 06/25/2019
• 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:
  US16/269,655
• Publication Number:
  US 20190169605A1
• Time in Patent Office: 138 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 inducible nuclease editing comprising:

a receptacle configured to receive cells;

one or more receptacles configured to receive nucleic acids, wherein the nucleic acids comprise an inducible nuclease editing system;

a growth module for growing cells for transformation;

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

a nuclease-directed editing module configured to activate the inducible nuclease editing system and 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 cells from the cell receptacle to the growth module, from the growth module to the transformation module and from the transformation 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 nucleic acids in the one or more receptacles comprise a backbone and an editing cassette, the automated multi-module cell editing instrument further comprises a nucleic acid assembly module, and the automated liquid handling system transfers the nucleic acids from the nucleic acids from the nucleic acid receptacle, from the nucleic acid receptacle to the nucleic acid assembly module, and from the nucleic acid assembly module to the transformation module.

3. The automated stand-alone multi-module cell editing instrument of claim 2, wherein the nucleic acid assembly module is configured to perform isothermal nucleic acid assembly.

4. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the automated liquid handling system comprises a sipper or pipettor.

5. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the inducible nuclease editing system is a chemically-induced inducible nuclease system.

6. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the inducible nuclease editing system is a virally-induced inducible nuclease system.

7. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the inducible nuclease editing system is a light-induced inducible nuclease system.

8. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the inducible nuclease editing system is a temperature-induced inducible nuclease system.

9. The automated stand-alone multi-module cell editing instrument of claim 8, wherein the inducible nuclease editing system is a heat-induced inducible nuclease system.

10. The automated stand-alone multi-module cell editing instrument of claim 1, wherein the receptacle configured to receive cells and the one or more receptacles configured to receive nucleic acids are configured to be contained within a reagent cartridge.

11. The automated stand-alone multi-module cell editing instrument of claim 10, wherein some or all reagents required for cell editing are received by the reagent cartridge.

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

a receptacle configured to receive cells;

a nucleic acid assembly module configured to assemble a vector backbone and an editing cassette, wherein the editing cassette comprises one or more inducible components of the inducible editing system, and wherein the nucleic acid assembly module is configured to accept and assemble nucleic acids to facilitate the desired genome editing events in the cells;

a growth module configured to grow the cells;

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

a nuclease-directed editing module configured to activate the inducible nuclease editing system and allow the assembled 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 script; and

an automated liquid handling system to move liquids from the cell receptacle to the growth module, from the growth module to the transformation module, and from the transformation module to the nuclease-directed editing module, as well as from the nucleic acid assembly module to the transformation module, all without user intervention.

13. The automated stand-alone multi-module cell editing instrument of claim 12, wherein the inducible nuclease editing system is a chemically-induced inducible nuclease system.

14. The automated stand-alone multi-module cell editing instrument of claim 12, wherein the inducible nuclease editing system is a virally-induced inducible nuclease system.

15. The automated stand-alone multi-module cell editing instrument of claim 12, wherein the inducible nuclease editing system is a light-induced inducible nuclease system.

16. The automated stand-alone multi-module cell editing instrument of claim 12, wherein the inducible nuclease editing system is a temperature-induced inducible nuclease system.

17. The automated stand-alone multi-module cell editing instrument of claim 16, wherein the inducible nuclease editing system is a heat-induced inducible nuclease system.

18. The automated stand-alone multi-module cell editing instrument of claim 12, further comprising at least one reagent cartridge containing reagents to perform inducible cell editing in the automated multi-module cell editing instrument.

19. The automated stand-alone multi-module cell editing instrument of claim 18, wherein the receptacles for the cells and nucleic acids are disposed within the reagent cartridge.

20. The automated stand-alone multi-module cell editing instrument of claim 18, wherein some or all reagents required for inducible cell editing are received by the reagent cartridge.