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Kristin M Whitworth, Joshua A Benne, Lee D Spate, Stephanie L Murphy, Melissa S Samuel, Clifton N Murphy, Jürgen A Richt, Eric Walters, Randall S Prather, Kevin D Wells
The CRISPR/Cas9 genome editing tool has increased the efficiency of creating genetically modified pigs for use as biomedical or agricultural models. The objectives were to determine if DNA editing resulted in a delay in development to the blastocyst stage or in a skewing of the sex ratio. Six DNA templates (gBlocks) that were designed to express guide RNAs that target the transmembrane protease, serine S1, member 2 (TMPRSS2) gene were in vitro transcribed. Pairs of CRISPR guide RNAs that flanked the start codon and polyadenylated Cas9 were co-injected into the cytoplasm of zygotes and cultured in vitro to the blastocyst stage...
October 15, 2016: Transgenic Research
Izumi Oda-Ishii, Atsushi Kubo, Willi Kari, Nobuhiro Suzuki, Ute Rothbächer, Yutaka Satou
[This corrects the article DOI: 10.1371/journal.pgen.1006045.].
October 2016: PLoS Genetics
Diting Qian, Zhiling Li, Yuting Zhang, Yue Huang, Que Wu, Gaizhen Ru, Man Chen, Bin Wang
Our study aimed to develop embryo models to evaluate the impact of oxidative stress on embryo development. Mouse zygotes, which stayed at G1 phase, were treated with prepared culture medium (containing 0.00, 0.01, 0.02, 0.03, 0.04, 0.05, or 0.1 mM hydrogen peroxide (H2O2)) for 30 min in experiment 1. The dose-effects of H2O2 on embryo development were investigated via comparisons of the formation rate at each stage (2- and 4-cell embryos and blastocysts). Experiment 2 was carried out to compare behaviors of embryos in a mild oxidative-stressed status (0...
2016: Oxidative Medicine and Cellular Longevity
Hong-Hao Yu, Heng Zhao, Yu-Bo Qing, Wei-Rong Pan, Bao-Yu Jia, Hong-Ye Zhao, Xing-Xu Huang, Hong-Jiang Wei
Dystrophinopathy, including Duchenne muscle dystrophy (DMD) and Becker muscle dystrophy (BMD) is an incurable X-linked hereditary muscle dystrophy caused by a mutation in the DMD gene in coding dystrophin. Advances in further understanding DMD/BMD for therapy are expected. Studies on mdx mice and dogs with muscle dystrophy provide limited insight into DMD disease mechanisms and therapeutic testing because of the different pathological manifestations. Miniature pigs share similar physiology and anatomy with humans and are thus an excellent animal model of human disease...
October 9, 2016: International Journal of Molecular Sciences
Yasir Ahmed-Braimah
Understanding the genetic basis of speciation is a central problem in evolutionary biology. Studies of reproductive isolation have provided several insights into the genetic causes of speciation, especially in taxa that lend themselves to detailed genetic scrutiny. Reproductive barriers have usually been divided into those that occur before zygote formation (prezygotic) and after (postzygotic), with the latter receiving a great deal of attention over several decades. Reproductive barriers that occur after mating but before zygote formation (postmating prezygotic) are especially understudied at the genetic level...
October 11, 2016: G3: Genes—Genomes—Genetics
Samantha L P Schilit, Masato Ohtsuka, Rolen M Quadros, Channabasavaiah B Gurumurthy
Microinjection of DNA expression cassettes into fertilized zygotes has been a standard method for generating transgenic animal models. While efficient, the injected DNA integrates randomly into the genome, leading to potential disruption of endogenous genes or regulatory elements, variation in copy number, or integration into heterochromatic regions that inhibit transgene expression. A recently developed method addresses such pitfalls of traditional transgenesis by targeting the transgene to predetermined sites in the genome that can safely harbor exogenous DNA...
October 11, 2016: Current Protocols in Human Genetics
Yue Zhang, Wei Li, Yongshun Ma, Dian Wang, Xiaoxue Zhao, Changjun Zeng, Ming Zhang, Xianyin Zeng, Qinggang Meng, Guangbin Zhou
The study was aimed to investigate the effect of melatonin on the development potential of mouse MII oocytes after cryopreservation. Mouse MII oocytes were subjected first to vitrification/warming and 2 h of in vitro culture (phase 1), then to parthenogenetic activation (PA) followed by in vitro culture of parthenogenetic embryos (phase 2). Different concentrations of melatonin (0, 10(-9), 10(-6) mol/L) were added to the medium during either phase 1, phase 2 or both phases. The fresh oocytes were used as control...
October 7, 2016: Cryobiology
Maella Severino-Freire, Nathalie Jonca, Melanie Pichery, Emilie Tournier, Nicolas Chassaing, Juliette Mazereeuw-Hautier
is missing (short communication).
October 10, 2016: Acta Dermato-venereologica
Miho M Suzuki, Tomoko Mori, Noriyuki Satoh
The initiation of embryonic gene expression in ascidian embryos appears to be tightly regulated by the number of DNA replication cycles. DNA methylation is thought to contribute to the clock mechanism that counts the rounds of DNA replication. We used mass spectrometry and whole genome bisulfite sequencing to characterize DNA methylation changes that occur in early developmental stages of the ascidian, Ciona intestinalis. We found that global DNA methylation in early Ciona development was static, and a base-wise comparison between the genomes of consecutive developmental stages found no DNA demethylation that was related to zygotic gene activation...
October 5, 2016: Genomics
Anne-Cecile Reymann, Fabio Staniscia, Anna Erzberger, Guillaume Salbreux, Stephan W Grill
Cytokinesis in eukaryotic cells is often accompanied by actomyosin cortical flow. Over 30 years ago, Borisy and White proposed that cortical flow converging upon the cell equator compresses the actomyosin network to mechanically align actin filaments. However, actin filaments also align via search-and-capture, and to what extent compression by flow or active alignment drive furrow formation remains unclear. Here we quantify the dynamical organization of actin filaments at the onset of ring assembly in the C...
October 10, 2016: ELife
Cuiqing Zhong, Jinsong Li
Haploid embryonic stem cells can be derived from androgenetic embryos produced by injection of sperm into enucleated oocytes or by removal of the female pronucleus from zygotes. These cells, termed AG-haESCs, can be used in place of sperm to produce the so-called semi-cloned (SC) mice. Importantly, AG-haESCs carrying H19-DMR and IG-DMR knockouts (DKO-AG-haESCs) can efficiently and stably support the generation of SC mice via intracytoplasmic AG-haESCs injection (ICAHCI), which provides a new route to obtain genetically modified mice...
2017: Methods in Molecular Biology
Sylvia J Bedford-Guaus, François Chauvigné, Eva Mejía-Ramírez, Mercè Martí, Antoni Ventura-Rubio, Ángel Raya, Joan Cerdà, Anna Veiga
While vitrification has become the method of choice for preservation of human oocytes and embryos, cryopreservation of complex tissues and of large yolk-containing cells, remains largely unsuccessful. One critical step in such instances is appropriate permeation while avoiding potentially toxic concentrations of cryoprotectants. Permeation of water and small non-charged solutes, such as those used as cryoprotectants, occurs largely through membrane channel proteins termed aquaporins (AQPs). Substitution of a Thr by an Ala residue in the pore-forming motif of the zebrafish (Dario rerio) Aqp3b paralog resulted in a mutant (DrAqp3b-T85A) that when expressed in Xenopus or porcine oocytes increased their permeability to ethylene glycol at pH 7...
October 5, 2016: Zygote: the Biology of Gametes and Early Embryos
A Mishra, I J Reddy, Psp Gupta, S Mondal
The objective of this study was to find out the impact of L-carnitine (10 mM) on developmental regulation of preimplantation sheep embryos cultured in vitro when supplemented in maturation medium and post-fertilization medium separately. Subsequent objective was to observe the L-carnitine-mediated alteration in expression of apoptotic genes (Bcl2, Bax, Casp3 and PCNA) in sheep oocytes and developing embryos produced in vitro. Oocytes matured with L-carnitine showed significantly (p < .05) higher cleavage (67...
September 30, 2016: Reproduction in Domestic Animals, Zuchthygiene
Yong Soo Hur, Eun Kyung Ryu, San Hyun Yoon, Kyung Sil Lim, Won Don Lee, Jin Ho Lim
OBJECTIVE: This study was conducted to compare the effects of static culture, dynamic culture, and the combination of dynamic culture with specialized surfaces involving co-culture on human embryonic development. Embryos cultured using conventional static culture (SC) techniques served as a control group. We compared dynamic culture using micro-vibration culture (MVC) and micro-vibration with co-culture (MCoC), in which autologous cumulus cells were used as a specialized surface. METHODS: We conducted a chart review of patients who were treated between January 2011 and November 2014 in order to compare embryonic development rates and pregnancy rates among the groups...
September 2016: Clinical and Experimental Reproductive Medicine
Tao Zhang, Yajun Yin, Huan Liu, Weili Du, Chonghua Ren, Ling Wang, Hongzhao Lu, Zhiying Zhang
CRISPR/Cas9 system has become a new versatile technology for genome engineering in various species. To achieve targeted modifications at the same site in both human and mice genomes by a CRISPR/Cas9 nuclease, we designed two target sites in conserved regions of vitamin D receptor (VDR) gene, which cover more than 17 kb of chromosome region depending on the species. We first validated the efficacy of single sgRNA mediated gene specific modifications were 36% and 31% in HEK293T cells. Concurrently, targeted of the intervening genomic segments deletions were generated in chromosomes when two sgRNAs worked simultaneously...
2016: PloS One
Manuel Adrian Troncoso-Ponce, Guillaume Barthole, Geoffrey Tremblais, Alexandra To, Martine Miquel, Loic Lepiniec, Sébastien Baud
In angiosperms, double fertilization of the embryo sac initiates the development of the embryo and the endosperm. In Arabidopsis thaliana, an exalbuminous species, the endosperm is reduced to one cell layer during seed maturation and reserves such as oil are massively deposited in the enlarging embryo. Here, we consider the strikingly different fatty acid (FA) compositions of the oils stored in the two zygotic tissues. Endosperm oil is enriched in ω-7 monounsaturated FAs, that represent more than 20 Mol% of total FAs, whereas these molecular species are ten-fold less abundant in the embryo...
September 28, 2016: Plant Cell
Karen B Barnard-Kubow, Nina So, Laura F Galloway
Genetic incompatibility is a hallmark of speciation. Cytonuclear incompatibilities are proposed to be among the first genetic barriers to arise during speciation. Accordingly, reproductive isolation within species should be heavily influenced by interactions between the organelle and nuclear genomes. However, there are few clear examples of cytonuclear incompatibility within a species. Here we show substantial postzygotic reproductive isolation in first-generation hybrids between differentiated populations of an herbaceous plant (up to 92% reduction in fitness)...
September 28, 2016: Evolution; International Journal of Organic Evolution
Dan-Dan Guo, Wen-Zhi Guan, Yi-Wen Sun, Jie Chen, Xia-Yun Jiang, Shu-Ming Zou
Fibroblast growth factor 1 (Fgf1) is known as a mitogenic factor involved in the regulation of cell growth, proliferation, and differentiation in vertebrates. Here, we report the isolation and characterization of two fgf1 genes in grass carp (Ctenopharyngodon idella). Grass carp fgf1a and fgf1b cDNAs are highly divergent, sharing a relatively low amino acid sequence identity of 50%, probably due to fish-specific gene duplication. fgf1a and fgf1b mRNAs were detected in the zygote and expressed throughout embryogenesis...
September 24, 2016: General and Comparative Endocrinology
Michael L Brosnahan, Lourdes Velo-Suárez, David K Ralston, Sophia E Fox, Taylor R Sehein, Alexi Shalapyonok, Heidi M Sosik, Robert J Olson, Donald M Anderson
Transitions between life cycle stages by the harmful dinoflagellate Alexandrium fundyense are critical for the initiation and termination of its blooms. To quantify these transitions in a single population, an Imaging FlowCytobot (IFCB), was deployed in Salt Pond (Eastham, Massachusetts), a small, tidally flushed kettle pond that hosts near annual, localized A. fundyense blooms. Machine-based image classifiers differentiating A. fundyense life cycle stages were developed and results were compared to manually corrected IFCB samples, manual microscopy-based estimates of A...
November 2015: Limnology and Oceanography
Adelita D Mendoza, Teresa K Woodruff, Sarah M Wignall, Thomas V O'Halloran
Zinc is an essential metal that serves as a cofactor in a variety of cellular processes, including meiotic maturation. Cellular control of zinc uptake, availability and efflux is closely linked to meiotic progression in rodent and primate reproduction where large fluctuations in zinc levels are critical at several steps in the oocyte-to-embryo transition. Despite these well-documented roles of zinc fluxes during meiosis, only a few of the genes encoding key zinc receptors, membrane-spanning transporters, and downstream signaling pathway factors have been identified to date...
September 21, 2016: Comparative Biochemistry and Physiology. Toxicology & Pharmacology: CBP
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