Use of haploid genomes for genetic diagnosis, modification and multiplication

Abstract

Methods for propagating haploid genomes of male or female origina and genetic screening and modification thereof are provided. These haploid genomes may be used to produce haploid embryos, and embryonic stem-like cells and differentiated cells. Also, these haploid genomes and cells containing, may be used as nuclear transfer donors to produce diploid nuclear transfer units. These diploid NT units e.g., human NT units, may be used to obtain pluripotent cells and differentiated cells and tissues.

1 . A method for selecting a haploid genome containing cell comprising the following steps: 
 (i) obtaining and amplifying the number of cells containing a haploid genome of male or female origin;    (ii) subjecting the genome of said haploid cells to genetic screening or analysis to determine whether said haploid genome comprises a desired genetic makeup; and    (iii) selecting cells that contain said desired genetic makeup.    
     
     
         2 . The method of  claim 1 , wherein said female-derived haploid cells are produced by activation of an oocyte in which half of the chromosomes are extruded in the polar body.  
     
     
         3 . The method of  claim 1 , wherein said female-derived haploid cells are produced by fertilization of an egg and removal of a male pronucleus therefrom.  
     
     
         4 . The method of  claim 1 , wherein said female-derived haploid cells are produced by activation of an egg to provide an egg containing two female pronuclei and removal of one of said pronuclei.  
     
     
         5 . The method of  claim 1 , wherein said female-derived haploid cells are produced by insertion of a diploid cell nucleus into an immature oocyte followed by separation of said chromosomes in to two haploid nuclei.  
     
     
         6 . The method of  claim 1 , wherein said female-derived haploid cells are produced by transfer of the nucleus of a parthenogenetic embryo (contains half the chromosomes) but propagated with the full DNA content (four chromatids) into an oocyte, and subsequent extrusion of half the chromosomes therefrom.  
     
     
         7 - 9 . (canceled)  
     
     
         10 . The method of  claim 1 , wherein said female-or male-derived haploid cells are amplified by a method selected from the group consisting of (i) allowing a haploid egg cytoplast to undergo cell division; (ii) allowing a haploid cell to produce a haploid embryo which is then cultured to produce “propagating haploid” cells; (iii) culturing a haploid embryo to produce propagating haploid cells and allowing such embryonic stem-like cells to differentiate; and (iv) culturing a haploid somatic cell cytoplast under conditions that allow cell division.  
     
     
         11 - 12 . (canceled)  
     
     
         13 . The method of  claim 1 , which further comprises using said selected male or female haploid genome or a cell containing said selected haploid genome is used to produce a diploid embryo.  
     
     
         14 . The method of  claim 1  which further comprises using a selected male and female haploid genome to produce a diploid embryo.  
     
     
         15 . The method of  claim 1 , wherein said selected male or female haploid genome or a cell containing said male or female genome is used as a nuclear transfer donor.  
     
     
         16 . The method of  claim 15 , wherein both a selected male or female haploid genome are used as nuclear transfer donors to produce a diploid nuclear transfer unit that contains said selected male and female haploid genome.  
     
     
         17 . The method of  claim 15 , wherein said haploid genome is human.  
     
     
         18 . The method of  claim 15 , wherein said haploid cell or genome comprises a differentiated cell embryonic stem-like cell or inner cell mass cell derived from a propagated haploid embryo.  
     
     
         19 . The method of  claim 18 , wherein said differentiated cells or embryonic stem-like cells are produced by in vitro culturing of a haploid embryo.  
     
     
         20 - 33 . (canceled)  
     
     
         34 . A propagating haploid cell line developed using the method of  claim 10 .  
     
     
         35 . The propagating haploid cell line of  claim 34 , wherein the cell line is a female cell-or male cell-derived haploid cell line.  
     
     
         36 . A diploid embryo that is produced by nuclear transfer wherein said nuclear transfer process comprises using as the nuclear transfer donor a haploid genome or cell containing which is produced according to  claim 1 .  
     
     
         37 . The diploid embryo of  claim 36  wherein said nuclear transfer process comprises transplantation of both a male and female haploid genome or cell containing produced according to  claim 1 .  
     
     
         38 . Embryonic stem-like cells or differentiated cells produced from a haploid embryo.  
     
     
         39 . An improved nuclear transfer process that is used to produce a cloned embryo, fetus or animal wherein the improvement comprises using as the nuclear transfer donor embryonic stem-like cells or differentiated cells produced from a haploid embryo.