Genetic markers and uses therefor

Abstract

The present invention relates, inter alia, to methods for determining whether or not an animal and/or its offspring is more likely than not to have an increased tolerance to heat, an increased resistance to ticks, and/or a desirable coat texture. It also provides methods for selecting or rejecting animals, one or more cells or embryos, estimating the worth of an animal, generating animals having a desired genotype/phenotype, cloning and breeding animals and herd formation.

1 . A method of selecting a variation in a prolactin receptor (PRLR) gene that results in a desirable phenotype in an animal that is a non-human mammal or an animal of family Phasiadnidae, comprising:
 (a) obtaining a nucleic acid sample from one or a plurality of the animals, or from one or a plurality of embryos or cells of the animal; and   (b) detecting in the nucleic acid sample of (a) at least one of:
 (i) one or more genetic variation in the PRLR gene, and 
 (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, and therefrom selecting the variation in the PRLR gene that results in the desirable phenotype, 
   wherein the desirable phenotype in the animal that is a non-human mammal comprises at least one of an increased tolerance to heat, an increased resistance to ticks, and a desirable coat texture which comprises at least one of a thin coat, a light coat, a sleek coat, and a short coat, relative, respectively, to the tolerance to heat, the resistance to ticks, and the coat texture in a non-human mammal that does not carry the one or more genetic variation,   and wherein the desirable phenotype in the animal of family Phasiadnidae comprises a desirable coat texture which comprises at least one of thin feathering and light feathering, relative to the coat texture in an animal of family Phasiadnidae that does not carry the one or more genetic variation.   
     
     
         2 . The method of  claim 1 , wherein the one or more genetic variation in the PRLR gene results in an increase in PRLR activity in the animal relative to PRLR activity in an animal that does not carry the one or more genetic variation. 
     
     
         3 . The method of  claim 1 , wherein the genetic variation in the PRLR gene is one or more of:
 (a) a variation resulting in a truncated PRLR polypeptide;   (b) a variation which results in truncation of a PRLR polypeptide in a region from approximately an amino acid position corresponding to position 430 to approximately an amino acid corresponding to position 490 of  Bos taurus  PRLR;   (c) a variation located in a final exon of the PRLR gene;   (d) a variation located within a region bounded by nucleotides corresponding to positions 39136469 and 39136649 of chromosome 20 of  Bos taurus;      (e) a variation at a nucleotide position corresponding to position 39136559 of chromosome 20 of  Bos taurus;      (f) a variation corresponding to a deletion of a C at a nucleotide position corresponding to position 39136559 of chromosome 20 of  Bos taurus ; and   (g) one or more genetic marker in linkage disequilibrium with at least one of (a)-(f).   
     
     
         4 . The method of  claim 1  which further comprises selecting the animal or the embryo or the cell of the animal of (a) based on its being more likely than not to have or result in the desirable phenotype, or rejecting the animal or the embryo or the cell of the animal of (a), based on its not being more likely than not to have or result in the desirable phenotype, said method comprising at least one of:
 (c) selecting the animal, embryo, or cell from which the nucleic acid sample has been obtained, which animal, embryo, or cell has been determined to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, wherein said genetic variation in the PRLR gene results in the desirable phenotype, and thereby selecting the animal, embryo, or cell that is more likely than not to have or result in the desirable phenotype; and 
 (d) rejecting the animal, embryo, or cell from which the nucleic acid sample has been obtained, which animal, embryo, or cell has been determined not to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, wherein said genetic variation in the PRLR gene results in the desirable phenotype, and thereby rejecting the animal, embryo, or cell that is not more likely than not to have or result in the desirable phenotype. 
 
     
     
         5 . The method of  claim 1  which further comprises calculating an estimated economic value or worth of the animal from which the nucleic acid sample has been obtained, based on one of (i) said one or more genetic variation in the PRLR gene, or (ii) said one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene as detected in step (b). 
     
     
         6 . The method of  claim 1  which further comprises breeding animals having the desirable phenotype, said method comprising:
 (c) selecting as a first animal the animal of (a) from which the nucleic acid sample has been obtained, which first animal has been determined to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene; and 
 (d) mating the first animal of (c) to a second animal, wherein the second animal is a non-human mammal or an animal of family Phasiadnidae selected from:
 (i) a randomly selected second animal, and 
 (ii) a second animal which has been determined, according to the method of claim  37 , to carry at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, and thereby breeding animals having the desirable phenotype. 
 
 
     
     
         7 . The method of  claim 1  which further comprises breeding an animal having the desirable phenotype, said method comprising:
 (c) selecting the embryo of (a) from which the nucleic acid sample has been obtained, which embryo has been determined to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene; and 
 (d) transferring the embryo to a gestational carrier, and thereby breeding an animal having the desirable phenotype. 
 
     
     
         8 . The method of  claim 1  which further comprises cloning an animal having the desirable phenotype, said method comprising:
 (c) selecting the embryo of (a) from which the nucleic acid sample has been obtained, which embryo has been determined to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, to obtain a selected embryo; and 
 (d) splitting by an embryo splitting technique the selected embryo of (c) to obtain a part of an embryo that is capable of being transferred to a gestational carrier, and thereby cloning an animal having the desirable phenotype. 
 
     
     
         9 . The method of  claim 1  which further comprises:
 (c) selecting the plurality of animals from which the nucleic acid sample has been obtained, which animals have been determined to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, thereby to obtain selected animals; 
 (d) rejecting the plurality of animals from which the nucleic acid sample has been obtained, which animals have been determined not to carry said at least one of (i) one or more genetic variation in the PRLR gene, and (ii) one or more genetic marker in linkage disequilibrium with one or more genetic variation in the PRLR gene, thereby to obtain rejected animals; and 
 (e) forming a herd of the selected animals of (c) and excluding from the herd the rejected animals of (d), thereby to form a herd of animals more likely than not to have the desirable phenotype. 
 
     
     
         10 . A method of producing a non-human mammalian cell or embryo capable of generating a non-human mammal which is more likely than not to have an increased tolerance to heat, an increased resistance to ticks, and/or a desirable coat texture, the method comprising altering the genomic DNA of a non-human mammalian cell or embryo which results in:
 an increase in prolactin receptor (PRLR) activity relative to the PRLR activity of a non-modified cell or embryo;   a truncation of the transcript or protein expressed from the PRLR gene;   a truncation of the transcript or protein expressed from the PRLR gene in a region from approximately an amino acid corresponding to position 430 to approximately an amino acid corresponding to position 490 of the  Bos taurus  PRLR;   a genetic alteration located in a final exon of the PRLR gene;   a genetic alteration located within a region bounded by nucleotides corresponding to positions 39136469 and 39136649 of chromosome 20 of  Bos taurus;      a genetic alteration in the nucleotide at a position corresponding to position 39136559 of chromosome 20 of  Bos taurus ; and/or   a genetic alteration corresponding to deletion of a cytosine (C) at a position corresponding to position 39136559 of chromosome 20 of  Bos taurus;      thereby producing a cell or embryo capable of generating a non-human mammal which is more likely than not to have an increased tolerance to heat, an increased resistance to ticks, and/or a desirable coat texture which comprises at least one of a thin coat, a light coat, a sleek coat, and a short coat, relative, respectively, to the tolerance to heat, the resistance to ticks, and the coat texture in animal non-human mammal that does not carry the one or more genetic variation.   
     
     
         11 . The method according to  claim 10 , wherein the cell is an oocyte or zygote. 
     
     
         12 . The method according to  claim 10 , wherein the step of altering the genomic DNA of the non-human mammalian cell or embryo is performed using a gene-editing technology. 
     
     
         13 . The method according to  claim 12 , wherein the gene-editing technology is selected from CRISPR/CAS, zinc finger nuclease, meganuclease or TALEN technology. 
     
     
         14 . The method according to  claim 10 , wherein the alteration is heterozygous. 
     
     
         15 . The method according to  claim 10 , wherein the alteration is homozygous. 
     
     
         16 . A method for generating a non-human mammal which is more likely than not to have an increased tolerance to heat, an increased resistance to ticks, and/or a desirable coat texture, the method comprising:
 (a) altering the genomic DNA of a non-human mammalian cell or embryo which results in:   an increase in prolactin receptor (PRLR) activity relative to the PRLR activity of a non-modified cell or embryo;   a truncation of the transcript or protein expressed from the PRLR gene;   a truncation of the transcript or protein expressed from the PRLR gene in a region from approximately an amino acid corresponding to position 430 to approximately an amino acid corresponding to position 490 of the  Bos taurus  PRLR;   a genetic alteration located in a final exon of the PRLR gene;   a genetic alteration located within a region bounded by nucleotides corresponding to positions 39136469 and 39136649 of chromosome 20 of  Bos taurus;      a genetic alteration in the nucleotide at a position corresponding to position 39136559 of chromosome 20 of  Bos taurus ; and/or   a genetic alteration corresponding to deletion of a cytosine (C) at a position corresponding to position 39136559 of chromosome 20 of  Bos taurus ; and   (b) generating a non-human mammal using the cell or embryo altered in step (a) which mammal is more likely than not to have an increased tolerance to heat, an increased resistance to ticks, and/or a desirable coat texture which comprises at least one of a thin coat, a light coat, a sleek coat, and a short coat, relative, respectively, to the tolerance to heat, the resistance to ticks, and the coat texture in animal non-human mammal that does not carry the one or more genetic variation.   
     
     
         17 . The method according to  claim 16 , wherein the cell is an oocyte or zygote. 
     
     
         18 . The method according to  claim 16 , wherein the step of altering the genomic DNA of the non-human mammalian cell or embryo is performed using a gene-editing technology. 
     
     
         19 . The method according to  claim 16 , wherein the gene-editing technology is selected from CRISPR/CAS, zinc finger nuclease, meganuclease or TALEN technology. 
     
     
         20 . The method according to  claim 16 , wherein the alteration is heterozygous. 
     
     
         21 . The method according to  claim 16 , wherein the alteration is homozygous. 
     
     
         22 . The method according to  claim 16 , wherein the non-human mammalian cell is an oocyte and the oocyte is fertilized using artificial insemination or in vitro fertilization. 
     
     
         23 . The method according to  claim 16 , wherein the non-human mammal is generated by transferring an embryo altered in step (a) to a gestational carrier. 
     
     
         24 . The method according to  claim 16 , wherein the non-human mammal is generated by a cloning process using a cell altered in step (a). 
     
     
         25 . A non-human mammal generated by the method of  claim 18 , comprising a gene-edited PRLR allele.