AUTOMATED CELL PROCESSING METHODS, MODULES, INSTRUMENTS, AND SYSTEMS
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.
1 . An automated stand-alone multi-module cell editing instrument for performing RNA-guided nuclease-directed editing in target regions in cells comprising:
a housing configured to house all or some of the modules;
a receptacle configured to receive the cells;
one or more receptacles configured to receive expression vectors, wherein the expression vectors introduce a desired DNA change to the target region and remove a proto-spacer motif (PAM) region from the target region;
a growth module for growing the cells for transfection;
a transformation module configured to introduce the expression vectors into the cells;
an RNA-guided nuclease-directed editing module configured to allow the introduced expression vectors to edit nucleic acids in the target region in the cells;
a selection module configured to select cells with a desired edit using the RNA-guided nuclease complexed with an gRNA complementary to the target region of the edited 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 cell receptacle to the growth module, from the growth module to the transformation module, from the transformation module to the RNA-guided nuclease-directed editing module, and from the RNA-guided nuclease-directed editing module to the selection module without user intervention.
2 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the RNA-guided nuclease is MAD7 nuclease.
3 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the RNA-guided nuclease-directed editing module is also a recovery module.
4 . The automated stand-alone multi-module cell editing instrument of claim 1 , further comprising a recovery module separate from the RNA-guided nuclease-directed editing module.
5 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the growth module is separate from the transformation module.
6 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the growth module and the transformation module are a same module.
7 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the liquid handling system comprises a pipettor.
8 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the selection module is a same module as the RNA-guided nuclease-directed editing module.
9 . The automated stand-alone multi-module cell editing instrument of claim 1 , comprising the selection module separate from the RNA-guided nuclease-directed editing module.
10 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the transfection module provides conditions for viral transduction.
11 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the cells are mammalian cells.
12 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the growth module and the RNA-guided nuclease-directed editing module are a same module.
13 . The automated stand-alone multi-module cell editing instrument of claim 1 , wherein the growth module is separate from the RNA-guided nuclease-directed editing module.
14 . An automated stand-alone multi-module cell editing instrument for performing RNA-guided nuclease-directed editing in target regions in mammalian cells comprising:
a housing configured to house all or some of the modules;
a receptacle configured to receive the mammalian cells;
one or more receptacles configured to receive RNA-guided nucleases and guide RNAs, wherein the guide RNAs introduce a desired DNA change to the target region and remove the proto-spacer motif (PAM) region from the target region;
a growth module for growing the mammalian cells for transfection;
a transfection module configured to introduce the RNA-guided nucleases and guide RNAs into the mammalian cells;
an RNA-guided nuclease-directed editing module configured to allow the introduced expression vectors to edit nucleic acids in the target region in the mammalian cells;
a selection module configured to select cells with a desired edit using the RNA-guided nuclease complexed with an gRNA complementary to the target region of the edited mammalian 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 cell receptacle to the growth module, from the growth module to the transfection module and from the transfection module to the RNA-guided nuclease-directed editing module without user intervention.
15 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the RNA-guided nuclease is MAD7 nuclease.
16 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the RNA-guided nuclease-directed editing module is a same module as the growth module.
17 . The automated stand-alone multi-module cell editing instrument of claim 14 , further comprising the RNA-guided nuclease-directed editing module separate from the growth module.
18 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the growth module is separate from the transfection module.
19 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the growth module and the transfection module are a same module.
20 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the liquid handling system comprises a pipettor.
21 . The automated stand-alone multi-module cell editing instrument of claim 20 , wherein the selection module is a same module as the RNA-guided nuclease-directed editing module.
22 . The automated stand-alone multi-module cell editing instrument of claim 14 , comprising the selection module separate from the RNA-guided nuclease-directed editing module.
23 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the receptacle configured to receive the mammalian cells is a same module as the growth module.
24 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the transfection module provides conditions for lipofection.
25 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the transfection module provides conditions for magnetofection.
26 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the transfection module provides conditions for transduction.
27 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the mammalian cells are human cells.
28 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the mammalian cells are mouse cells.
29 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the mammalian cells are rat cells.
30 . The automated stand-alone multi-module cell editing instrument of claim 14 , wherein the receptacle configured to receive the mammalian cells is separate from the growth module.