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Juanjuan Xu, Rui Fang, Li Chen, Daozhen Chen, Jian-Ping Xiao, Weimin Yang, Honghua Wang, Xiaoqing Song, Ting Ma, Shiping Bo, Chong Shi, Jun Ren, Lei Huang, Li-Yi Cai, Bing Yao, X Sunney Xie, Sijia Lu
Preimplantation genetic screening (PGS) is widely used to select in vitro-fertilized embryos free of chromosomal abnormalities and to improve the clinical outcome of in vitro fertilization (IVF). A disadvantage of PGS is that it requires biopsy of the preimplantation human embryo, which can limit the clinical applicability of PGS due to the invasiveness and complexity of the process. Here, we present and validate a noninvasive chromosome screening (NICS) method based on sequencing the genomic DNA secreted into the culture medium from the human blastocyst...
September 29, 2016: Proceedings of the National Academy of Sciences of the United States of America
Stefan C Kane, Elissa Willats, Sammya Bezerra Maia E Holanda Moura, Jonathan Hyett, Fabrício da Silva Costa
Chromosomal aneuploidy is responsible for a significant proportion of pregnancy failures, whether conceived naturally or through in vitro fertilization (IVF). In an effort to improve the success rate of IVF, screening embryos for aneuploidy - or pre-implantation genetic screening (PGS) - has been proposed as a means of ensuring only euploid embryos are selected for transfer. Early PGS approaches were based on fluorescence in situ hybridization testing, and have been shown not to improve live birth rates. Recent developments in genetic testing technologies - such as next-generation sequencing and quantitative polymerase chain reaction, coupled with embryo biopsy at the blastocyst stage - have shown promise in improving IVF outcomes, but they remain to be validated in adequately powered, prospective randomized trials...
September 29, 2016: Fetal Diagnosis and Therapy
Mousa I Shamonki, Helen Jin, Zachary Haimowitz, Lian Liu
OBJECTIVE: To assess whether preimplantation genetic screening (PGS) is possible by testing for free embryonic DNA in spent IVF media from embryos undergoing trophectoderm biopsy. DESIGN: Prospective cohort analysis. SETTING: Academic fertility center. PATIENT(S): Seven patients undergoing IVF and 57 embryos undergoing trophectoderm biopsy for PGS. INTERVENTION(S): On day 3 of development, each embryo was placed in a separate media droplet...
August 23, 2016: Fertility and Sterility
Gheona Altarescu
Abstract During the last two decades prenatal genetic screening and diagnosis has become the cornerstone of medical care for family planning to prevent genetic disease. Carrier screening programs for genetic disorders that are prevalent in various populations identify couples and pregnancies at risk of having an affected child. These couples can proceed with a choice of invasive prenatal diagnosis tests of the fetus (chorionic villous sampling and amniocentesis), or non-invasive prenatal testing of free fetal DNA circulation in the maternal blood which has emerged within the last few years and is currently available for fetal sexing for X Linked disorders...
June 2016: Pediatric Endocrinology Reviews: PER
Stephanie G Valderramos, Rashmi R Rao, Emily W Scibetta, Neil S Silverman, Christina S Han, Lawrence D Platt
BACKGROUND: Since its commercial release in 2011 cell-free DNA screening has been rapidly adopted as a routine prenatal genetic test. However, little is known about its performance in actual clinical practice. OBJECTIVE: We sought to investigate factors associated with the accuracy of abnormal autosomal cell-free DNA results. STUDY DESIGN: We conducted a retrospective cohort study of 121 patients with abnormal cell-free DNA results from a referral maternal-fetal medicine practice from March 2013 through July 2015...
June 28, 2016: American Journal of Obstetrics and Gynecology
Naoki Takeda, Kazuya Yoshinaga, Kenryo Furushima, Kazufumi Takamune, Zhenghua Li, Shin-Ichi Abe, Shin-Ichi Aizawa, Ken-Ichi Yamamura
Protamines are expressed in the spermatid nucleus and allow denser packaging of DNA compared with histones. Disruption of the coding sequence of one allele of either protamine 1 (Prm1) or Prm2 results in failure to produce offspring, although sperm with disrupted Prm1 or Prm2 alleles are produced. Here, we produced Prm1-deficient female chimeric mice carrying Prm1-deficient oocytes. These mice successfully produced Prm1(+/-) male mice. Healthy Prm1(+/-) offspring were then produced by transferring blastocysts obtained via in vitro fertilization using zona-free oocytes and sperm from Prm1(+/-) mice...
2016: Scientific Reports
Theofania Bounartzi, Konstantinos Dafopoulos, George Anifandis, Christina I Messini, Chrysoula Koutsonikou, Spyros Kouris, Maria Satra, Sotirios Sotiriou, Nicholas Vamvakopoulos, Ioannis E Messinis
The purpose of this study was to evaluate the predictive value of free sperm plasma DNA (f-spDNA) and sperm DNA fragmentation (SDF), in semen specimens from men undergoing in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET) treatments. Fifty-five semen samples were evaluated during 55 consecutive IVF/ICSI-ET cycles. F-spDNA was determined by conventional quantitative real-time PCR-Sybr green detection approach, while evaluation of sperm DNA damage was performed using the sperm chromatin dispersion (SCD) assay...
April 2016: Human Fertility: Journal of the British Fertility Society
L Sarno, R Revello, E Hanson, R Akolekar, K H Nicolaides
OBJECTIVES: First, to examine in twin pregnancies the performance of first-trimester screening for fetal trisomies 21, 18 and 13 by cell-free (cf) DNA testing of maternal blood and, second, to compare twin and singleton pregnancies regarding the distribution of fetal fraction of cfDNA and rate of failure to obtain a result. METHODS: This was a prospective study in 438 twin and 10 698 singleton pregnancies undergoing screening for fetal trisomies by cfDNA testing at 10 + 0 to 13 + 6 weeks' gestation...
June 2016: Ultrasound in Obstetrics & Gynecology
Takayuki Sakurai, Akiko Kamiyoshi, Hisaka Kawate, Chie Mori, Satoshi Watanabe, Megumu Tanaka, Ryuichi Uetake, Masahiro Sato, Takayuki Shindo
The CRISPR/Cas9 system is capable of editing multiple genes through one-step zygote injection. The preexisting method is largely based on the co-injection of Cas9 DNA (or mRNA) and guide RNAs (gRNAs); however, it is unclear how many genes can be simultaneously edited by this method, and a reliable means to generate transgenic (Tg) animals with multiple gene editing has yet to be developed. Here, we employed non-inheritable maternal Cas9 (maCas9) protein derived from Tg mice with systemic Cas9 overexpression (Cas9 mice)...
2016: Scientific Reports
Hong-Cui Zhao, Ting Ding, Yun Ren, Tian-Jie Li, Rong Li, Yong Fan, Jie Yan, Yue Zhao, Mo Li, Yang Yu, Jie Qiao
STUDY QUESTION: Does Sirt3 dysfunction result in poor developmental outcomes for human oocytes after in vitro maturation (IVM)? SUMMARY ANSWER: Inefficient Sirt3 expression induced decreased mitochondrial DNA copy number and biogenesis, and therefore impaired the developmental competence of human IVM oocytes. WHAT IS KNOWN ALREADY: Cytoplasmic immaturity in IVM oocytes may lead to reduced developmental competence. Mitochondrial dysfunction results in the accumulation of free radicals and leads to DNA mutations, protein damage, telomere shortening and apoptosis...
March 2016: Human Reproduction
Renee Stokowski, Eric Wang, Karen White, Annette Batey, Bo Jacobsson, Herb Brar, Madhumitha Balanarasimha, Desiree Hollemon, Andrew Sparks, Kypros Nicolaides, Thomas J Musci
OBJECTIVE: To evaluate the clinical performance of non-invasive prenatal testing for trisomy 21, 18, and 13 using targeted cell-free DNA (cfDNA) analysis. METHODS: Targeted cfDNA analysis using DANSR™ and FORTE™ with microarray quantitation was used to evaluate the risk of trisomy 21, 18, and 13 in blinded samples from 799 singleton, twin, natural, and IVF pregnancies. Subjects either had fetal chromosome evaluation by karyotype, FISH, QF-PCR, or karyotype for newborns with suspected aneuploidy at birth...
December 2015: Prenatal Diagnosis
Sabine Traver, Elodie Scalici, Tiffany Mullet, Nicolas Molinari, Claire Vincens, Tal Anahory, Samir Hamamah
Cell-free DNA (cfDNA) fragments, detected in blood and in other biological fluids, are released from apoptotic and/or necrotic cells. CfDNA is currently used as biomarker for the detection of many diseases such as some cancers and gynecological and obstetrics disorders. In this study, we investigated if cfDNA levels in follicular fluid (FF) samples from in vitro fertilization (IVF) patients, could be related to their ovarian reserve status, controlled ovarian stimulation (COS) protocols and IVF outcomes. Therefore, 117 FF samples were collected from women (n = 117) undergoing IVF/Intra-cytoplasmic sperm injection (ICSI) procedure and cfDNA concentration was quantified by ALU-quantitative PCR...
2015: PloS One
Bedri Karakas, Wafa Qubbaj, Saad Al-Hassan, Serdar Coskun
The discovery of cell-free fetal DNA (cfDNA) circulating in the maternal blood has provided new opportunities for noninvasive prenatal diagnosis (NIPD). However, the extremely low levels of cfDNA within a high background of the maternal DNA in maternal circulation necessitate highly sensitive molecular techniques for its reliable use in NIPD. In this proof of principle study, we evaluated the earliest possible detection of cfDNA in the maternal plasma by a bead-based emulsion PCR technology known as BEAMing (beads, emulsion, amplification, magnetics)...
2015: PloS One
Naresh L Selokar, Monika Saini, Himanshu Agrawal, Prabhat Palta, Manmohan S Chauhan, Radheysham Manik, Suresh K Singla
Aberrant epigenetic reprogramming, especially genomic hypermethylation, is implicated as the primary reason behind the failure of the cloning process during somatic cell nuclear transfer (SCNT). We transfected one-cell-stage zona-free buffalo embryos produced by handmade cloning with 50 nM DNMT1 small interfering RNA (siRNA), using lipofectamine, to knockdown the DNA methyltransferase 1 (DNMT1) gene. siRNA treatment decreased (p<0.001) the expression level of DNMT1 mRNA and DNMT1 protein in the one-cell-stage embryos and increased (p<0...
April 2015: Cellular Reprogramming
Haitao Wu, Chenhui Ding, Xiaoting Shen, Jing Wang, Rong Li, Bing Cai, Yanwen Xu, Yiping Zhong, Canquan Zhou
To develop a noninvasive medium-based preimplantation genetic diagnosis (PGD) test for α-thalassemias-SEA. The embryos of α-thalassemia-SEA carriers undergoing in vitro fertilization (IVF) were cultured. Single cells were biopsied from blastomeres and subjected to fluorescent gap polymerase chain reaction (PCR) analysis; the spent culture media that contained embryo genomic DNA and corresponding blastocysts as verification were subjected to quantitative-PCR (Q-PCR) detection of α-thalassemia-SEA. The diagnosis efficiency and allele dropout (ADO) ratio were calculated, and the cell-free DNA concentration was quantitatively assessed in the culture medium...
March 2015: Medicine (Baltimore)
Yueqiu Tan, Xuyang Yin, Shuoping Zhang, Hui Jiang, Ke Tan, Jian Li, Bo Xiong, Fei Gong, Chunlei Zhang, Xiaoyu Pan, Fang Chen, Shengpei Chen, Chun Gong, Changfu Lu, Keli Luo, Yifan Gu, Xiuqing Zhang, Wei Wang, Xun Xu, Gábor Vajta, Lars Bolund, Huanming Yang, Guangxiu Lu, Yutao Du, Ge Lin
BACKGROUND: Next generation sequencing (NGS) is now being used for detecting chromosomal abnormalities in blastocyst trophectoderm (TE) cells from in vitro fertilized embryos. However, few data are available regarding the clinical outcome, which provides vital reference for further application of the methodology. Here, we present a clinical evaluation of NGS-based preimplantation genetic diagnosis/screening (PGD/PGS) compared with single nucleotide polymorphism (SNP) array-based PGD/PGS as a control...
2014: GigaScience
Jason M Franasiak, Xinxin Yao, Elena Ashkinadze, Todd Rosen, Richard T Scott
BACKGROUND: Multimodal prenatal screening for developmental pathology is increasingly common. In this case, definitive prenatal diagnosis of androgen insensitivity syndrome was diagnosed after discordant results from karyotypes determined by embryonic preimplantation genetic screening and antenatal ultrasound results. CASE: A 38-year-old woman, gravida 2 para 0010, undergoing in vitro fertilization with preimplantation genetic screening transferred one male and one female embryo...
February 2015: Obstetrics and Gynecology
E Bevilacqua, M M Gil, K H Nicolaides, E Ordoñez, V Cirigliano, H Dierickx, P J Willems, J C Jani
OBJECTIVES: To report clinical implementation of cell-free DNA (cfDNA) analysis of maternal blood in screening for trisomies 21, 18 and 13 in twin pregnancies and examine variables that could influence the failure rate of the test. METHODS: cfDNA testing was performed in 515 twin pregnancies at 10-28 weeks' gestation. The failure rate of the test to provide results was compared with that in 1847 singleton pregnancies, and logistic regression analysis was used to determine which factors among maternal and pregnancy characteristics were significant predictors of test failure...
January 2015: Ultrasound in Obstetrics & Gynecology
E Scalici, S Traver, N Molinari, T Mullet, M Monforte, E Vintejoux, S Hamamah
STUDY QUESTION: Could cell-free DNA (cfDNA) quantification in individual human follicular fluid (FF) samples become a new non-invasive predictive biomarker for in vitro fertilization (IVF) outcomes? SUMMARY ANSWER: CfDNA level in human follicular fluid samples was significantly correlated with embryo quality and could be used as an innovative non-invasive biomarker to improve IVF outcomes. WHAT IS KNOWN ALREADY: CfDNA fragments, resulting from apoptotic or necrotic events, are present in the bloodstream and their quantification is already used as a biomarker for gynaecological and pregnancy disorders...
December 2014: Human Reproduction
Yuta Ishizuka, Toru Takeo, Satohiro Nakao, Hidetaka Yoshimoto, Yumiko Hirose, Yuki Sakai, Yuka Horikoshi, Shiori Takeuji, Shuuji Tsuchiyama, Naomi Nakagata
Hyaluronidase is generally used to remove cumulus cells from mouse oocytes before oocyte cryopreservation, intracytoplasmic sperm injection or DNA injection. In general, use of cumulus-free mouse oocytes decreases in vitro fertilizing ability compared with cumulus-surrounded oocytes. The effect of hyaluronidase exposure on the quality of mouse oocytes is not fully understood. Here, we investigated the effect of hyaluronidase exposure time on the fertilization rate of fresh and vitrified mouse oocytes and their subsequent developmental ability in vitro...
2014: Journal of Reproduction and Development
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