# US 10,765,084 B2 · Method for improving the mean dry shoot weight, mean dry grain weight, and suppressing seed-borne infection in a cereal crop > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,765,084 B2 - Document ID: US10765084B2 - Status: Granted - Kind: B2 - Application number: 15/507,585 - Filed: 2015-08-29 - Published: 2017-09-28 - Granted: 2020-09-08 - Earliest claimed: 2014-08-29 - Assignees: The Provost, Fellows, Foundation Scholars, and the Other Members of Board, of the College of the Holy and Undivided Trinity of Queen Elizabeth Near Dublin; University College Dublin, National University of Ireland, Dublin - Applicants: The Provost, Fellows, Scholars and other members of Board of Trinity College Dublin; University College Dublin, National University of Ireland, Dublin - Inventors: Brian Murphy; Trevor Hodkinson; Fiona Doohan - Web page: https://patentgrep.com/patent/US10765084B2 ## CPC - A01N 63/30: Microbial fungi; Substances produced thereby or obtained therefrom - A01C 1/06: Coating or dressing seed - A01C 1/08: Immunising seed - A01H 17/00: Symbiotic or parasitic combinations including one or more new plants, e.g. mycorrhiza (lichens A01H15/00) - A01N 63/36: Penicillium - C12N 15/82: for plant cells {, e.g. plant artificial chromosomes (PACs)} - A01N: PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF (preservation of food or foodstuff A23); BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES (preparations for medical, dental or toiletry purposes which kill or prevent the growth or proliferation of unwanted organisms A61K); PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS - A01C: PLANTING; SOWING; FERTILISING (parts, details or accessories of agricultural machines or implements, in general A01B51/00 - A01B75/00) - A01H: NEW PLANTS OR {NON-TRANSGENIC} PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract A seed coating composition comprising a mixture of a fungal root endophyte isolated from a root of the plant found in low-nutrient, drought-stressed or multiply-stressed conditions and a carrier medium for application of said composition to a target seed, wherein the endophyte is characterised in having a nuclear ribosomal internal transcribed spacer (nr ITS) with at least a 90% sequence identity to any one of SEQ ID NOs: 3 to 15. ## Claims ### Claim 1 A seed coating composition for use in (a) increasing grain yield and dry shoot weight in a plant, the composition comprising a mixture of a fungal root endophyte isolated from a root of the plant and a carrier medium for application of said composition to a target seed, or for (b) increasing tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds, the composition comprising a mixture of a fungal root endophyte isolated from a root of a plant sprouted from said seeds and a carrier medium for application of said composition to a target seed, wherein the endophyte is characterised in having a nuclear ribosomal internal transcribed spacer (nrITS) with 100% sequence identity to any one of SEQ ID NOs: 3 to 15, wherein the fungal root endophyte is isolated from a root of the plant Hordeum murinum, and wherein the plant and seeds are in low-nutrient conditions, or in drought-stressed conditions, or in multiply-stressed conditions. ### Claim 2 (depends on 1) A seed coating composition according to claim 1, wherein the plant is Hordeum murinum subsp. murinum L. (Hmm). ### Claim 3 (depends on 1) The seed coating composition according to claim 1, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof. ### Claim 4 (depends on 1) The seed coating composition according to claim 1, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the composition for increasing tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds planted in low-nutrient, drought-stressed or multiply-stressed conditions comprises the fungal root endophytes having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6 or SEQ ID NO: 11. ### Claim 5 (depends on 1) The seed coating composition according to claim 1, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the composition for increasing grain yield and dry shoot weight in a plant found in low-nutrient, drought-stressed or multiply-stressed conditions comprises at least one of the fungal endophyte isolates having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 7 or SEQ ID NO: 8. ### Claim 6 (depends on 1) The seed coating composition according to claim 1, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the composition for increasing tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds planted in low-nutrient, drought-stressed or multiply-stressed conditions comprises the fungal root endophytes having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 13, SEQ ID NO: 14 or SEQ ID NO: 15. ### Claim 7 (depends on 1) The seed coating composition according to claim 1, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the composition for increasing grain yield and dry shoot weight in a plant found in low-nutrient, drought-stressed or multiply-stressed conditions comprises at least one of the fungal endophyte isolates having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 13, SEQ ID NO: 14 OR SEQ ID NO: 15. ### Claim 8 (depends on 1) The seed coating composition according to claim 1, wherein the concentration of the fungal endophyte isolate in the composition is between 0.001% (w/v) to 1.0% (w/v). ### Claim 9 (depends on 1) The seed coating composition according to claim 1, wherein the carrier medium is selected from water, distilled water, sterilised water, sterilised distilled water, an emulsified suspension, wettable powder, encapsulation of spores of the fungal endophyte in alginate beads, or a film-coating. ### Claim 10 (depends on 1) A seed coated with, mixed with or encapsulated with the composition of any one of claim 1. ### Claim 11 (depends on 1) A seed mixed with the composition according to claim 1, in which the carrier medium is alginate beads. ### Claim 12 A method of preparing a seed having (a) increased grain yield and dry shoot weight or (b) with increased tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds, the method comprising the step of inoculating the seed with a fungal root endophyte isolated from a root of a plant which produced said seed, wherein the endophyte is characterised in having a nuclear ribosomal internal transcribed spacer (nrITS) with 100% sequence identity to any one of SEQ ID NOs: 3 to 15, wherein the fungal root endophyte is isolated from a root of the plant Hordeum murinum, and wherein the plant and seeds are in low-nutrient conditions, or in drought-stressed conditions, or in multiply-stressed conditions. ### Claim 13 (depends on 12) The method according to claim 12, wherein the plant is Hordeum murinum subsp. murinum L. (Hmm). ### Claim 14 (depends on 12) The method according to claim 12, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof. ### Claim 15 (depends on 12) The method according to claim 12, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the method for preparing a seed with increased tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds comprises inoculating the seed with a fungal root endophyte having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6 or SEQ ID NO: 11, or variants thereof. ### Claim 16 (depends on 12) The method according to claim 12, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the method for increasing grain yield and dry shoot weight in a plant comprises inoculating the seed with at least one of the fungal endophyte isolates having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 7 or SEQ ID NO: 8. ### Claim 17 (depends on 12) The method according to claim 12, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the method for preparing a seed with increased tolerance to the development of seed-borne fungal infections on germinated and ungerminated seeds comprises inoculating the seed with a fungal root endophyte having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 13, SEQ ID NO: 14 or SEQ ID NO: 15. ### Claim 18 (depends on 12) A method according to claim 12, wherein the fungal root endophyte is selected based on having a nuclear ribosomal internal transcribed spacer selected from any one of SEQ ID NOs: 3 to 15, or any combination thereof, and wherein the method for increasing grain yield and dry shoot weight in a plant comprises inoculating the seed with at least one of the fungal endophyte isolates having a nuclear ribosomal internal transcribed spacer as defined by SEQ ID NO: 6, SEQ ID NO: 8, SEQ ID NO: 13, SEQ ID NO: 14 OR SEQ ID NO: 15. ### Claim 19 (depends on 12) The method according to claim 12, wherein the concentration of the fungal endophyte isolate is between 0.001% (w/v) to 1.0% (w/v). ### Claim 20 (depends on 12) The method according to claim 12, wherein the carrier medium is selected from water, distilled water, sterilised water, sterilised distilled water, an emulsified suspension, wettable powder, encapsulation of spores of the fungal endophyte in alginate beads, or a film-coating.