# US 2024/0200005 A1 · Method for developing and/or reprogramming plant cellular objects > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 2024/0200005 A1 - Document ID: US20240200005A1 - Status: Published application - Kind: A1 - Application number: 18/555,347 - Filed: 2022-04-14 - Published: 2024-06-20 - Earliest claimed: 2021-04-15 - Applicants: Screensys GmbH - Inventors: Jialan CAO; Alexander GROSS; Johann Michael KÖHLER; Patrick SCHAUB; Klaus PALME; Oleksandr DOVZHENKO - Web page: https://patentgrep.com/patent/US20240200005A1 ## CPC - B01L 3/0203: {Burettes, i.e. for withdrawing and redistributing liquids through different conduits} - G01N 15/1409: Handling samples, e.g. injecting samples - B01L 3/0262: {using touch-off at substrate or container} - B01L 3/502761: {specially adapted for handling suspended solids or molecules independently from the bulk fluid flow, e.g. for trapping or sorting beads or physically stretching molecules} - B01L 3/502784: {specially adapted for droplet or plug flow, e.g. digital microfluidics} - C12M 23/16: {Microfluidic devices; Capillary tubes (integrated microfluidic structures B01L3/5027; microreactors B01J19/0093)} - C12M 41/46: {of cellular or enzymatic activity or functionality, e.g. cell viability} - C12M 47/04: {Cell isolation or sorting (purging biological preparations of unwanted cells C12N5/0081, determining the presence or kind of microorganism C12Q1/04)} - C12N 5/04: Plant cells or tissues {(culture media C12N5/0025)} - G01N 15/1434: Optical arrangements - G01N 15/1484: {microstructural devices} - G01N 15/149: specially adapted for sorting particles, e.g. by their size or optical properties - G01N 33/0098: {Plants or trees (wood G01N33/46)} - G01N 35/08: using a stream of discrete samples flowing along a tube system, e.g. flow injection analysis - B01L 2200/0647: Handling flowable solids, e.g. microscopic beads, cells, particles - B01L 2200/0673: Handling of plugs of fluid surrounded by immiscible fluid - B01L 2300/0861: Configuration of multiple channels and/or chambers in a single devices - B01L 2400/0487: fluid pressure, pneumatics - B01L 2400/082: Active control of flow resistance, e.g. flow controllers - C12N 2523/00: Culture process characterised by temperature - C12N 2529/10: Stimulation by light - G01N 2015/1006: {for cytology} - G01N 2035/00237: {Handling microquantities of analyte, e.g. microvalves, capillary networks} - B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE - G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q) - C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS} - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract The present invention relates to a method for developing and/or reprogramming plant cellular objects comprising the steps: providing a reservoir containing a medium with plant cellular objects: providing a first set of compartments of sample fluid embedded in carrier fluid in a microfluidic conduit, wherein the carrier fluid is immiscible with the medium, wherein the first set's compartments of sample fluid each comprise medium and at least one plant cellular object: providing one or more first state triggers to the plant cellular objects in the microfluidic conduit for inducing a first state in the plant cellular objects of the first set of compartments: incubating the plant cellular objects of the first set of compartments in the microfluidic conduit for a time span sufficient for the plant cellular objects to transfer to the first state: selecting one or more first selection parameters indicative of the first state: identifying, within the first set of compartments in the microfluidic conduit, compartments according to the one or more first selection parameters and optionally assigning the compartments with respective state identifiers. ## Claims ### Claim 1 A method for developing and/or reprogramming plant cellular objects comprising the steps: a) providing a reservoir containing a medium with plant cellular objects; b) providing a first set of compartments of sample fluid embedded in carrier fluid in a microfluidic conduit, wherein the carrier fluid is immiscible with the medium, wherein the first set's compartments of sample fluid each comprise medium and at least one plant cellular object; c) providing one or more first state triggers to the plant cellular objects in the microfluidic conduit for inducing a first state in the plant cellular objects of the first set of compartments; d) incubating the plant cellular objects of the first set of compartments in the microfluidic conduit for a time span sufficient for the plant cellular objects to transfer to the first state; e) selecting one or more first selection parameters indicative of the first state; f) identifying, within the first set of compartments in the microfluidic conduit, compartments according to the one or more first selection parameters and optionally assigning the compartments with respective state identifiers. ### Claim 2 (depends on 1) The method of claim 1, further comprising the step: g) sorting the first set's compartments according to their state as identified in step f), e.g. separating the compartments identified in step f) as comprising plant cellular objects that have, according to the one or more first selection parameters, transferred to the first state from the compartments that were not identified in step f) as comprising plant cellular objects that have transferred to the first state. ### Claim 3 (depends on 1) The method of claim 1, further comprising: h) providing a second set of compartments of sample fluid embedded in carrier fluid in the microfluidic conduit, wherein the second set's compartments of sample fluid each comprise medium and at least one plant cellular object; i) providing one or more second state triggers to the second set's plant cellular objects in the microfluidic conduit for inducing a second state in the plant cellular objects of the second set of compartments; j) incubating the plant cellular objects of the second set of compartments in the microfluidic conduit for a time span sufficient for the plant cellular objects to transfer to the second state; k) selecting one or more second selection parameters indicative of the second state; l) identifying, within the second set of compartments in the microfluidic conduit, compartments with plant cellular objects that have transferred to the second state and optionally assigning the compartments with respective state identifiers. ### Claim 4 (depends on 1) The method of claim 1, comprising: assigning a compartment identifier or a set identifier to the compartments. ### Claim 5 (depends on 1) The method of claim 1, further comprising the step: transferring the sample fluid compartments to a subsequent procedural step in accordance with one or a combination of: i. their compartment identifier; ii. their set identifier; iii. their state identifier. ### Claim 6 (depends on 1) The method of claim 1, wherein the sample fluid compartments or sets of sample fluid compartments are treated individually. ### Claim 7 (depends on 1) The method of claim 1, wherein the trigger(s) include(s): changing the composition of the sample fluid in the compartments as compared to the medium upon formation of the sample fluid compartments; or changing the composition of the sample fluid in the compartments after formation of the sample fluid compartments while the compartments are in the microfluidic channel. ### Claim 8 (depends on 7) The method of claim 7, wherein different sets of sample compartments are provided with different trigger substances. ### Claim 9 (depends on 7) The method of claim 7, wherein different compartments of sample fluid within the first or second set are provided with the same trigger substance at different concentrations. ### Claim 10 (depends on 1) The method of claim 1, wherein the state trigger comprises at least one substance selected from the group comprising culture medium salts or organic components, small chemical molecules, plant growth regulators or macromolecules, temperature and light regimes, or combinations of different triggers. ### Claim 11 (depends on 1) The method of claim 1, wherein the step of providing the state trigger(s), i.e. step(s) c)/i), include(s) subjecting at least some of the compartments or at least one of the sets of compartments to one or a combination of light shock, heat shock and cold shock conditions for inducing the respective state, wherein: the heat shock conditions include an elevated temperature of at least 5° C. above standard incubation temperature; the cold shock conditions include a lowered temperature of at least 5° C. or 10° ° C. below standard incubation temperature; and the light shock includes a change of the lighting conditions as compared to standard incubation. ### Claim 12 (depends on 1) The method of claim 1, wherein the microfluidic conduit is a tube that is at least 50 cm long. ### Claim 13 (depends on 1) The method of claim 1, wherein step e) includes: optically detecting the compartments while the compartments are in the tube, and creating imaging data thereby. ### Claim 14 (depends on 1) The method of claim 1 comprising: dispensing a plurality of plant cellular objects that are embedded in the compartments of sample fluid in accordance with the identifiers of the respective compartments. ### Claim 15 (depends on 1) The method of claim 1, wherein the carrier fluid is immiscible with the sample fluid(s). ### Claim 16 (depends on 2) The method of claim 2, wherein step g) comprises: sorting the first set's compartments according to their state identifiers. ### Claim 17 (depends on 3) The method of claim 3, further comprising the step: m) sorting the second set's compartments according to their state as identified in step l). ### Claim 18 (depends on 17) The method of claim 17, wherein step m) includes separating the compartments identified in step l) as comprising plant cellular objects that have, according to the one or more second selection parameters, transferred to the second state from the compartments that were not identified in step l) as comprising plant cellular objects that have transferred to the second state. ### Claim 19 (depends on 17) The method of claim 17, wherein step m) includes sorting the second set's compartments according to their state identifiers. ### Claim 20 (depends on 1) The method of claim 1, further comprising the step: discarding the compartments with a state identifier that indicates that the compartment was not identified as comprising plant cellular objects that have transferred to the respective state. ### Claim 21 (depends on 6) The method of claim 6, wherein the sample fluid compartments or sets of sample fluid compartments are treated according to their compartment identifier or set identifier or state identifier. ### Claim 22 (depends on 14) The method of claim 14, wherein the plant cellular objects of respective compartments are dispensed at different target sites in accordance with the identifiers of the respective compartments.