Described herein are methods of enhancing chromosomal homologous recombination to stimulate a loss of heterozygosity at a gene locus of interest in a living cell. These methods are driven by an enhancer component and a target-specific endonuclease component and proceed through a mechanism whereby: exogenous donor DNA that is homologous to the gene locus of interest is not introduced into the living cell; the desired allele of the gene locus of interest remains uncleaved; and the undesired allele is either uncleaved, cleaved at a single location, or cleaved at multiple locations. These methods have numerous applications, including the repair of risk alleles for disease prevention, the correction of heterozygous mutations in dividing cells, the design of cancer therapeutics, and the design of novel gene-drive strategies.

Methods of producing human stem cells are disclosed for parthenogenetically activating human oocytes by manipulation of O.sub.2 tension, including manipulation of Ca.sup.2+ under high O.sub.2 tension and contacting oocytes with serine threonine kinase inhibitors under low O.sub.2 tension, isolating inner cell masses (ICMs) from the activated oocytes, and culturing the cells of the isolated ICMs under high O.sub.2 tension. Moreover, methods are described for the production of stems cells from activated oocytes in the absence of non-human animal products, including the use of human feeder cells/products for culturing ICM/stem cells. Stem cells produced by the disclosed methods are also described.

Methods of producing human stem cells are disclosed for parthenogenetically activating human oocytes by manipulation of O.sub.2 tension, including manipulation of Ca.sup.2+ under high O.sub.2 tension and contacting oocytes with serine threonine kinase inhibitors under low O.sub.2 tension, isolating inner cell masses (ICMs) from the activated oocytes, and culturing the cells of the isolated ICMs under high O.sub.2 tension. Moreover, methods are described for the production of stems cells from activated oocytes in the absence of non-human animal products, including the use of human feeder cells/products for culturing ICM/stem cells. Stem cells produced by the disclosed methods are also described.

Methods of enhancing fertility of a female subject by increasing the number of oogonia present in the ovary of the female subject are provided. Aspects of the methods include methods of in vivo expansion of oogonia as well as methods of ex vivo expansion of oogonia.

The present invention provides modified oocytes having a nuclear genome derived from a first oocyte and cytoplasm derived from a second oocyte from a different subject, and methods for making and using such modified oocytes. The methods and compositions of the present invention can be useful in a variety of settings including, but not limited to, in in vitro fertilization (“IVF”) procedures.

The present invention provides modified oocytes having a nuclear genome derived from a first oocyte and cytoplasm derived from a second oocyte from a different subject, and methods for making and using such modified oocytes. The methods and compositions of the present invention can be useful in a variety of settings including, but not limited to, in in vitro fertilization (IVF) procedures.

Provided for the first time in the present invention is a method for preparing a non-human primate somatic cell cloned animal, which method specifically comprises the steps of: (i) providing a reconstructed egg, wherein the egg comes from the non-human primate (ii) activating the reconstructed egg to form an activated reconstructed egg or activated reconstructed embryo formed by the reconstructed egg; (iii) reprogramming (a) the activated reconstructed egg or (b) embryonic cells of the activated reconstructed embryo to obtain a reprogrammed reconstructed egg or reprogrammed reconstructed embryo; and (iv) regenerating the reprogrammed reconstructed egg or reprogrammed reconstructed embryo to obtain the non-human primate somatic cell cloned animal. The method of the present invention can significantly improve the developmental capacity of nucleus-transplanted embryos in non-human primates (such as monkeys).

The present invention provides methods and compositions to improve the efficiency of somatic cell nuclear transfer (SCNT) of human cells and the consequent production of human nuclear transfer ESC (hNT-ESCs). More specifically, the present invention relates to the discovery that trimethylation of Histone H3-Lysine 9 (H3K9me3) in reprogramming resistant regions (RRRs) in the nuclear genetic material of human donor somatic cells prevents efficient human somatic cell nuclear reprogramming or SCNT. The present invention provide methods and compositions to decrease H3K9me3 in methods to improve efficacy of hSCNT by exogenous or overexpression of the demethylase KDM4 family and/or inhibiting methylation of H3K9me3 by inhibiting the histone methyltransferases SUV39h1 and/or SUV39h2.

The present invention provides methods for making reconstructed diploid human oocytes comprising the diploid genome of a human somatic cell, and also methods for making human nuclear transfer embryos, human embryonic stem cells, and human differentiated cells therefrom. The present invention also provides reconstructed human oocytes, human nuclear transfer embryos, human embryonic stem cells, and differentiated cells made using such methods, as well as compositions and kits useful in performing such methods.

The present invention provides modified oocytes having a nuclear genome derived from a first oocyte and cytoplasm derived from a second oocyte from a different subject, and methods for making and using such modified oocytes. The methods and compositions of the present invention can be useful in a variety of settings including, but not limited to, in in vitro fertilization (IVF) procedures.