Revealing the bovine embryo transcript profiles during early in vivo embryonic development

Vallée, Maud; Dufort, Isabelle; Desrosiers, Stéphanie; Labbe, Aurélie; Gravel, Catherine; Gilbert, Isabelle; Robert, Claude; Sirard, Marc‐André

doi:10.1530/rep-08-0533

Cited by 30 publications

(20 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We obtained between 1.3 and 2.1 ng of total RNA per embryo (Figure 1). These amounts correspond closely to other reports using an identical methodology [19] and the same RNA extraction kit [20,21] used presently. Examination of the 18S and 28S rRNA fractions by micro-electrophoresis showed profiles similar to those previously observed in total RNA derived from bovine blastocysts [19,20] and possessing high RQI quality scores (>9).…”

Section: Resultssupporting

confidence: 88%

RNA-seq analysis of single bovine blastocysts

Chitwood

Rincón

Kaiser³

et al. 2013

BMC Genomics

View full text Add to dashboard Cite

BackgroundUse of RNA-Seq presents unique benefits in terms of gene expression analysis because of its wide dynamic range and ability to identify functional sequence variants. This technology provides the opportunity to assay the developing embryo, but the paucity of biological material available from individual embryos has made this a challenging prospect.ResultsWe report here the first application of RNA-Seq for the analysis of individual blastocyst gene expression, SNP detection, and characterization of allele specific expression (ASE). RNA was extracted from single bovine blastocysts (n = 5), amplified, and analyzed using high-throughput sequencing. Approximately 38 million sequencing reads were generated per embryo and 9,489 known bovine genes were found to be expressed, with a high correlation of expression levels between samples (r > 0.97). Transcriptomic data was analyzed to identify SNP in expressed genes, and individual SNP were examined to characterize allele specific expression. Expressed biallelic SNP variants with allelic imbalances were observed in 473 SNP, where one allele represented between 65-95% of a variant’s transcripts.ConclusionsThis study represents the first application of RNA-seq technology in single bovine embryos allowing a representation of the embryonic transcriptome and the analysis of transcript sequence variation to describe specific allele expression.

show abstract

Section: Resultssupporting

confidence: 88%

RNA-seq analysis of single bovine blastocysts

Chitwood

Rincón

Kaiser³

et al. 2013

BMC Genomics

View full text Add to dashboard Cite

show abstract

“…The Pico Pure RNA Isolation kit used in this study was specifically designed for low RNA content samples with low elution volumes. The use of this kit followed by DNase treatment during the RNA extraction has been reported in several gene expression studies of early embryos (Vallée et al, 2009;Larman et al, 2011). As also shown in this study, DNase I digestion is an effective method for removing DNA contamination from RNA samples, and hence produced efficient cDNA synthesis (Tavares et al, 2011).…”

Section: A B C Discussionsupporting

confidence: 54%

Effect of DNase treatment on RNA extraction from preimplantation murine embryos

et al. 2015

View full text Add to dashboard Cite

ABSTRACT. The quality of RNA is crucial when performing microarray experiments. This is particularly important when dealing with preimplantation embryos, from which a minimum yield of RNA of good quality can be produced. We report the optimization of several RNA extraction methods applied to preimplantation embryos at different stages of development. The quality of the samples was confirmed using a microarray and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) analysis. A total of 30 cultured two-cell stage embryos of ICR mice were pooled at the 8-cell, morula, and blastocyst stages. The embryos were divided into two groups comprising DNase- treated and non-DNase-treated RNA samples. Total RNA was extracted using a Pico Pure RNA Isolation Kit following the manufacturer protocol, with some modifications. Lysed samples were bound to a silica-based filter, treated with deoxyribonuclease I (DNase I), and washed several times before elution. RNA concentration and integrity were evaluated using an Agilent 2100 Bioanalyzer and an RNA 6000 Pico Assay kit. Although concentrations of non-DNase-treated RNAs were higher than DNase-treated RNA, DNase-treated RNA gave a higher RNA integrity number compared with non-DNase-treated RNA. Inclusion of DNase treatment in the RNA extraction procedure gave the best quality RNA samples from preimplantation embryos, as validated by microarray and RT-qPCR quality control.

show abstract

“…The aRNA concentration varied between 227.5 and 798.1 ng mL À1 . The amplification step was validated according to the methods described by Vallée et al (2009) and Gilbert et al (2010).…”

Section: Microarray-based Transcriptome Studymentioning

confidence: 99%

Insulin exposure during in vitro bovine oocyte maturation changes blastocyst gene expression and developmental potential

Laskowski

Humblot

Sirard

et al. 2017

Reprod. Fertil. Dev.

Self Cite

View full text Add to dashboard Cite

Abstract. Metabolic imbalance impairs fertility, because changes in concentrations of metabolites and hormones in the blood and follicular fluid create an unfavourable environment for early embryonic development. Insulin is a key metabolic hormone known for its effects on fertility: insulin concentrations are increased during energy balance disturbances in diabetes or metabolic syndrome. Still, insulin is frequently used at supraphysiological concentrations for embryo in vitro culture with unknown consequences for the developmental potential of the offspring. In the present study we investigated the effects of insulin exposure during in vitro bovine oocyte maturation on developmental rates, embryo quality and gene expression. Supplementation of the maturation media with insulin at 10 or 0.1 mg mL À1 decreased blastocyst rates compared with an insulin-free control (19.8 AE 1.3% and 20.4 AE 1.3% vs 23.8 AE 1.3%, respectively; P , 0.05) and led to increased cell numbers (nearly 10% more cells on Day 8 compared with control; P , 0.05). Transcriptome analysis revealed significant upregulation of genes involved in lipid metabolism, nuclear factor (erythroid-derived 2)-like 2 (NRF2) stress response and cell differentiation, validated by quantitative polymerase chain reaction. To conclude, the results of the present study demonstrate that insulin exposure during in vitro oocyte maturation has a lasting effect on the embryo until the blastocyst stage, with a potential negative effect in the form of specific gene expression perturbations.

show abstract

Revealing the bovine embryo transcript profiles during early in vivo embryonic development

Cited by 30 publications

References 40 publications

RNA-seq analysis of single bovine blastocysts

RNA-seq analysis of single bovine blastocysts

Effect of DNase treatment on RNA extraction from preimplantation murine embryos

Insulin exposure during in vitro bovine oocyte maturation changes blastocyst gene expression and developmental potential

Contact Info

Product

Resources

About