Project description:Oocyte maturation defects are one contributing factor in unexplained female infertility. Failure of some oocytes to undergo germinal vesicle breakdown or progress to second meiotic metaphase in response to an ovulatory stimulus can limit the number of high quality oocytes available for assisted reproduction. Understanding the molecular mechanisms that underlie oocyte maturation failure may lead to novel methods for increasing the number of high quality oocytes available for assisted reproduction, as well as providing new insight into the basic biology of oocyte maturation. Using RNA sequencing, we compared the transcriptome of rhesus monkey oocytes that failed to mature (FTM) in response to an ovulatory stimulus in vivo to normal germinal vesicle and metaphase II stage oocytes.
Project description:The goal of this study was to discover the underlying mechanisms that drive major changes in gene regulation during preimplantation development in the rhesus monkey. RNAseq data were collected from preimplantation stages (8-cell, morula, blastocyst), and used in conjunction with data from metaphase II stage oocytes to characterize individual gene expression profiles during preimplantation development, identify major groups of co-regulated genes, and identify likely upstream factors driving that gene regulation. Embryos were cultured following in vitro fertilization as described and lysed for analysis at specific stages.
Project description:Pre-implantation embryogenesis encompasses several critical events including genome reprogramming, zygotic genome activation (ZGA), and cell fate commitment, of which most remain mechanistically unclear in primate. Here we carried out time-series RNA-seq in 26 single and 8 pooled rhesus monkey oocytes and pre-implantation embryos encompassing representative developmental stages to explore these process.
Project description:Oocyte maturation defects are one contributing factor in unexplained female infertility. Failure of some oocytes to undergo germinal vesicle breakdown or progress to second meiotic metaphase in response to an ovulatory stimulus can limit the number of high quality oocytes available for assisted reproduction. Understanding the molecular mechanisms that underlie oocyte maturation failure may lead to novel methods for increasing the number of high quality oocytes available for assisted reproduction, as well as providing new insight into the basic biology of oocyte maturation. Using RNA sequencing, we compared the transcriptome of rhesus monkey oocytes that failed to mature (FTM) in response to an ovulatory stimulus in vivo to normal germinal vesicle and metaphase II stage oocytes.
Project description:Rhesus Monkey is one of the important primate models widely used in the fields of disease mechanism study, pre-clinical test in drug discovery and molecular evolution. However, the majority of rhesus gene annotations were putatively mapped from human genome, with only 10% supported by rhesus EST data.So, to better study the transcriptome, paired-end, strand-specific, poly(A)-positive RNA-Seq were performed in 5 rhesus monkey tissues. 5 tissue samples examined: prefrontal cortex, liver, skeletal muscle, adipose, testis
Project description:Derivation of embryonic stem cells (ESC) genetically identical to a patient by somatic cell nuclear transfer (SCNT) holds the potential to cure or alleviate the symptoms of many degenerative diseases while circumventing any immunorejection issues. However, no primate nuclear transfer embryonic stem (ntES) cell lines have been derived to date. Here, we used a modified SCNT technique to produce rhesus macaque SCNT blastocysts at a relatively high efficiency from adult donor cells and we successfully derived two primate ntES cell lines from 304 oocytes (an overall efficiency of 0.7%). Nuclear and mitochondrial DNA analysis confirmed the ntES cell lines were derived from rhesus monkey SCNT blastocysts and both rhesus monkey ntES cell lines exhibited a normal ESC morphology, expressed key stemness markers, were transcriptionally indistinguishable from control ESC lines and differentiated into multiple cell types. This is, to our knowledge, the first confirmed derivation of primate ntES cell lines. Keywords: rhesus monkey somatic cell nuclear transfer embryonic stem cells
Project description:Embryonic stem cells (ESCs) may be able to cure or alleviate the symptoms of various degenerative diseases. However, unresolved issues regarding apoptosis, maintaining function and tumor formation mean a prudent approach should be taken towards advancing ESCs into human clinical trials. The rhesus monkey provides the ideal model organism for developing strategies to prevent immune rejection and test the feasibility, safety and efficacy of ESC-based medical treatments. Transcriptional profiling of rhesus ESCs provides a foundation for future pre-clinical ESC research using non-human primates as the model organism. In this research we use microarray, immunocytochemistry, real-time and standard RT-PCR to characterize and transcriptionally profile rhesus monkey embryonic stem cells. We identify 367 rhesus monkey stemness genes, we demonstrate the high level (>85%) of conservation of rhesus monkey stemness gene expression across five different rhesus monkey embryonic stem cell lines, we demonstrate that rhesus monkey ESC lines maintain a pluripotent undifferentiated state over a wide range of Pou5f1 (Oct-4) expression levels and we compare rhesus monkey, human and murine stemness genes to identify the key mammalian stemness genes. The supplementary tables list the genes that have been upregulated in each undifferentiated rhesus monkey embryonic stem cell line (GSM99998, GSM99999,GSM100000, GSM100001, GSM100002, GSM99965, GSM99966) in comparison analysis with the pooled differentiated embryonic stem cells (GSM99840). Supplemental Table 1 contains the comparison analysis for all 52,865 probe sets on the rhesus monkey gene chip, Supplemental Table 2 contains the rhesus monkey genes that were significantly upregulated (FC>3) in the ORMES-6 biological replicates, Supplemental Table 3 contains the rhesus monkey genes that were significantly upregulated (FC>3) in the pooled differentiated EBs and Supplemental Tables 4-8 represent genes that were significantly upregulated in ORMES 6A, 7, 9, 10 and 13 respectively. Supplemental Table 9 contains the RT-PCR primers used in this project. Keywords: Rhesus monkey embryonic stem cell microarray