258 MESSENGER RNA EXPRESSION PATTERNS OF HISTONE MODIFICATION GENES IN BOVINE EMBRYOS DERIVED FROM DIFFERENT ORIGINS

2006 ◽  
Vol 18 (2) ◽  
pp. 236 ◽  
Author(s):  
M. Nowak-Imialek ◽  
C. Wrenzycki ◽  
D. Herrmann ◽  
I. Lagutina ◽  
A. Lucas-Hahn ◽  
...  
Keyword(s):  
Histone Deacetylase ◽  
Relative Abundance ◽  
Transcriptional Activation ◽  
Messenger Rna ◽  
Expression Patterns ◽  
Rna Expression ◽  
Bovine Embryos ◽  
Different Origins

Epigenetic modifications of the genome, such as covalent modifications of histones, are crucial for transcriptional regulation during development. The N-terminal tails of the histones are subject to post-translational modifications, including acetylation, deacetylation and methylation. histone acetylation loosens chromatin packing and correlates with transcriptional activation. The enzymes Histone acetyltransferases (HATs) transfer acetyl moieties to the lysine residues of histones H2A, H2B, H3, and H4. Histone acetylation is a reversible process which is catalyzed by the histone deacetylase (HDAC) and results in transcriptional repression. Histone methyltransferase (HMT) is responsible for the methylation of arginine in histones 3 and 4, playing an important role in transcriptional activation of genes. In contrast, the H3 Lys 9 methylation is associated with a transcriptionally repressive heterochromatin. The objective of the present study was to determine the effects of different origins of embryos on the relative abundance of transcripts for the histone acetyltransferase 1 (HAT1), histone deacetylase 2 (HDAC2), histone metyltransferases (SUV39H1 and G9A), and heterochromatin protein 1 (HP1). Messenger RNA expression profiles of these genes were investigated in bovine oocytes and pre-implantation embryos up to the blastocyt stage produced either in vitro by two different culture systems, i.e. SOF+BSA or TCM+SERUM, by somatic cloning using adult male and female fibroblasts, parthenogenetic activation, and androgenetic construction, or in vivo, employing semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Significant differences are described below. HAT1, SUV39H1, G9A, and HP1 mRNA transcripts decreased in enucleated oocytes, compared with immature oocytes. The relative abundance of HAT1 and SUV39H1 transcripts was significantly increased in NT-derived zygotes produced from adult female fibroblasts, compared to their in vitro fertilized and parthenogenetic counterparts. No differences were found in the relative abundances of gene transcripts at the 8-16-cell stage, except for parthenogenetic embryos in which SUV39H1 transcripts were significantly higher than in all other 8-16 cell groups. The relative abundance of SUV39H1, G9A, and HP1 transcripts were significantly higher in NT-derived blastocysts derived from adult male fibroblasts than in their in vivo-generated counterparts. HP1 and G9A transcript levels were significantly increased in NT-derived blastocysts derived from male fibroblasts compared to NT-derived embryos produced from female fibroblasts. The results show that the in vitro environment and the nuclear transfer protocol affect mRNA expression patterns of histone modification genes in pre-implantation bovine embryos.

247 MESSENGER RNA EXPRESSION PATTERNS OF DNA AND HISTONE METHYLTRANSFERASES IN PREIMPLANTATION DEVELOPMENT OF IN VIVO- AND IN VITRO-PRODUCED BOVINE EMBRYOS

2006 ◽  
Vol 18 (2) ◽  
pp. 231 ◽  
Author(s):  
K. Höffmann ◽  
H. Niemann ◽  
K.-G. Hadeler ◽  
D. Herrmann ◽  
C. Wrenzycki
Keyword(s):  
Developmental Stage ◽  
Mrna Expression ◽  
Relative Abundance ◽  
Expression Patterns ◽  
Bovine Embryos ◽  
Histone Methyltransferases ◽  
Embryonic Genome ◽  
Uterine Flushing

The effects of in vitro production (IVP) and/or somatic nuclear transfer on mRNA expression patterns have mostly been determined in morulae and blastocysts, i.e. after embryonic genome activation. Comparative data regarding mRNA expression patterns throughout the oviductal phase of pre-implantation development are scarce. Here we studied mRNA expression for genes related to DNA methylation and modification of histones which account for the major epigenetic reprogramming during development. Pertubated epigenetic reprogramming of the genome is a likely cause of developmental abnormalities and epigenetic diseases associated with assisted reproduction technologies. The objective of the present study was to compare mRNA expression of DNA methyltransferases Dnmt1, -3a, and -3b and histone methyltransferases SUV39-h1 and G9a between in vivo-derived bovine embryos and their IVP counterparts using a semiquantitative RT-PCR assay (Wrenzycki et al. 2002 Biol. Reprod. 66, 127-134) employing two embryos for each assay. In vivo-derived embryos were collected from 28 superovulated heifers by endoscopic flushing of oviducts (zygotes to 8-cell stages) (Besenfelder et al. 2001 Theriogenology 55, 837-845) or by uterine flushing (16-cell stages to blastocysts). Endoscopic flushing at different time points after AI (Days 1, 1.5, 2, 3, 4, and 4.5) yielded 31 zygotes; 15 two-cell, 5 three-cell, 13 four-cell, 1 five-cell, 2 six-cell, and 11 eight-cell embryos; 4 degenerated embryos; and 18 unfertilized ova. The recovery rate (corpora lutea counted per recovered embryos) was 58% and 62% for the endoscopic and uterine flushing, respectively. Differences in the relative abundance of each gene transcript between groups were tested using ANOVA with the main effects being origin (in vivo/in vitro) and developmental stage (zygote to blastocyst) and their interactions followed by multiple pairwise comparisons using a Tukey test (P < 0.05). Origin of embryos affected the relative abundance of transcripts for Dnmt1, Dnmt3a, and SUV39-h1, and developmental stage affected the relative abundance of transcripts for Dnmt1, -3a, -3b, SUV39-h1, and G9a. No interactive effects were observed for origin and developmental stage in the relative abundance of all transcripts. The relative abundance of Dnmt1 transcripts differed significantly between in vivo- and in vitro-produced morulae and blastocysts. For Dnmt3a, mRNA differences were determined between in vivo- and in vitro-produced 10-16-cell stages and morulae. Suv39-h1 transcripts differed significantly between in vivo- and in vitro-derived zygotes, 2-cell embryos, 8-cell embryos, 10-16-cell embryos, and blastocysts. The results suggest that IVP alters mRNA expression of genes related to epigenetic modifications very early in development, even before the embryonic genome has been activated.

scholarly journals Relative messenger RNA expression profiling of collagenases and aggrecanases in human articular chondrocytes in vivo and in vitro

2002 ◽  
Vol 46 (10) ◽  
pp. 2648-2657 ◽  
Author(s):  
Brigitte Bau ◽  
Pia M. Gebhard ◽  
Jochen Haag ◽  
Thomas Knorr ◽  
Eckart Bartnik ◽  
...  
Keyword(s):  
Expression Profiling ◽  
Messenger Rna ◽  
Articular Chondrocytes ◽  
Human Articular Chondrocytes ◽  
Rna Expression ◽  
Rna Expression Profiling

scholarly journals Evaluation of biomarkers for in vitro prediction of drug-induced nephrotoxicity in RPTEC/TERT1 cells

2020 ◽  
Vol 9 (2) ◽  
pp. 91-100 ◽  
Author(s):  
Xuan Qiu ◽  
Yufa Miao ◽  
Xingchao Geng ◽  
Xiaobing Zhou ◽  
Bo Li
Keyword(s):  
Transcriptional Activation ◽  
Messenger Rna ◽  
Aristolochic Acid ◽  
Kidney Injury ◽  
Drug Induced ◽  
Vitro Assay ◽  
Protein Levels ◽  
Aristolochic Acid I

Abstract There have been intensive efforts to identify in vivo biomarkers that can be used to monitor drug-induced kidney damage before significant impairment occurs. Kidney injury molecule-1, neutrophil gelatinase-associated lipocalin, clusterin, β2-microglobulin and cystatin C (CysC) have been validated as clinical or preclinical biomarkers in urinary and plasma predictive of acute and chronic kidney injuries and diseases. A high-throughput in vitro assay predictive of nephrotoxicity could potentially be implemented in early drug discovery stage to reduce attrition at later stages of drug development. To assess the potential of these known in vivo biomarkers for in vitro evaluation of drug-induced nephrotoxicity, we selected four nephrotoxic agents (cisplatin, cyclosporin, aristolochic acid I and gentamicin) and detected their effects on the protein levels of nephrotoxic biomarkers in RPTEC/TERT1 cells. The protein levels of clusterin, CysC, GSTπ and TIMP-1 significantly increased in the conditioned media of RPTEC/TERT1 cells treated with cisplatin, cyclosporin, aristolochic acid I and gentamicin. The messenger RNA levels of clusterin, CysC, GSTπ and TIMP-1 also increased in RPTEC/TERT1 cells treated with cisplatin, cyclosporin, aristolochic acid I and gentamicin, indicating that drug-induced upregulation involves transcriptional activation. Taken together, the results clearly demonstrate that among the known in vivo nephrotoxic biomarkers, clusterin, CysC, GSTπ and TIMP-1 can be effectively used as in vitro biomarkers for drug-induced nephrotoxicity in RPTEC/TERT1 cells.

Histone deacetylase–mediated transcriptional activation reduces proviral loads in HTLV-1–associated myelopathy/tropical spastic paraparesis patients

Blood ◽  
2007 ◽  
Vol 110 (10) ◽  
pp. 3722-3728 ◽  
Author(s):  
Agnès Lezin ◽  
Nicolas Gillet ◽  
Stéphane Olindo ◽  
Aïssatou Signaté ◽  
Nathalie Grandvaux ◽  
...  
Keyword(s):  
Histone Deacetylase ◽  
Transcriptional Activation ◽  
Gene Activation ◽  
Spastic Paraparesis ◽  
Viral Gene Expression ◽  
Tropical Spastic Paraparesis ◽  
Viral Reservoir ◽  
Viral Gene

AbstractEpigenetic modifications of chromatin may play a role in maintaining viral latency and thus persistence of the human T-lymphotropic virus type 1 (HTLV-1), which is responsible for HTLV-associated myelopathy/tropical spastic paraparesis (HAM/TSP). A major determinant of disease progression is increased peripheral blood proviral load (PVL), possibly via the accumulation of infected cells in the central nervous system (CNS) creating a damaging inflammatory response. Current therapeutic approaches that focus on reducing either cell proliferation, viral replication, or tissue invasion are still unsatisfactory. Contrasting with these inhibitory strategies, we evaluated the efficacy of a novel approach aimed, paradoxically, at activating viral gene expression to expose virus-positive cells to the host immune response. We used valproate (VPA), a histone deacetylase inhibitor that has been used for decades as a chronic, safe treatment for epileptic disorders. Based on in vitro and in vivo data, we provide evidence that transient activation of the latent viral reservoir causes its collapse, a process that may alleviate the condition of HAM/TSP. This represents the first such approach to treating HAM/TSP, using gene activation therapy to tilt the host-pathogen balance in favor of an existing antiviral response. This trial is registered at http://clinicaltrials.gov/as no. NCT00519181.

scholarly journals Targeting the Tumour-Specific Spliceosome Through In Silico Virtual Screening for Discovery of New SF3B1 Small Molecule Inhibitors

2018 ◽  
Vol 4 (Supplement 2) ◽  
pp. 201s-201s
Author(s):  
L.Z. Wong
Keyword(s):  
Virtual Screening ◽  
Vitamin D3 ◽  
Transcriptional Activation ◽  
Messenger Rna ◽  
Binding Free Energy ◽  
Specific Cell ◽  
Breast Cancers ◽  
Eukaryotic Genes

Background: The coding (exon) and noncoding (intron) eukaryotic genes are expressed as precursor messenger RNA (premRNA). mRNA splicing defines the process by which introns are excised from premRNA and flanking exons are ligated together. This process is catalyzed by the spliceosome in which combination of small nuclear ribonucleoproteins (U1, U2, U4, U5, U6) forms a spliceosome. The SF3B1 protein is a core component of the U2 snRNP that binds to the branch site and facilitate RNA splicing. Recent studies have identified mutations and dysregulation of in SF3B1 activities in subsets of human cancers including chronic lymphocytic leukemias (CLLs), uveal melanomas, pancreatic cancers and breast cancers. Despite promising in vivo results indicating the potential of spliceosome modulators in targeting the refractory breast cancers, the preclinical and clinical development of such modulators will take several years to complete. Aim: In the current study, we sought to identify new SF3B1 modulators using massive virtual screening of FDA-approved drugs or novel agents for drug repurposing. Methods: A total of 3000 compounds were screened and the hits were identified based on the binding free energy (kcal/mol) of the molecules to the predicted binding sites. Of the 90 hits, vitamin D3 and its analogs (calcipotriol and calcitriol) were identified as a putative SF3B1 modulators. Results: Further in vitro testing revealed that vitamin D3 and its analogs induced significant mRNA misplicing and tumor-specific cell death in MCF7 and MDA-MB-468. Further analyses revealed that vitamin D3 and its analogs significantly reduce SF3B1 protein expression with no changes in its mRNA expression. Conclusion: These results suggest that vitamin D3 and its analogs might interact with SF3B1 to induce protein degradation rather than transcriptional activation.

204 RELATIVE MESSENGER RNA ABUNDANCE OF HYALURONAN RECEPTORS AND SYNTHASES ON IN VITRO- AND IN VIVO-DERIVED DAY 7 AND DAY 13 BOVINE EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 200
Author(s):  
M. Clemente ◽  
A. T. Palasz ◽  
J. de La Fuente ◽  
P. Lonergan ◽  
A. Gutierrez-Adan ◽  
...  
Keyword(s):  
Embryo Development ◽  
Messenger Rna ◽  
Developmental Stages ◽  
Transcript Abundance ◽  
Early Embryo ◽  
Bovine Embryos ◽  
Cd44 Receptor ◽  
Mrna Extraction

Hyaluronan (HA), which progressively increases during embryogenesis, is a glycosaminoglycan that plays a major role in oocyte/embryo development (Fenderson et al. 1993 Differentiation 54, 85–95). One of the main functions of HA is to participate in the cell proliferation and migration that are controlled by HA receptors, RHAMM and C44, and by the presence of different HA synthases, Has1, Has2, and Has3. All have very distinctive features and functions at different stages of embryo development. The objective of this study was to determine the relative mRNA abundance of HA receptors and synthases in Day 7 and 13 bovine embryos derived in vitro or in vivo. In vitro embryos were produced by standard oocyte maturation and fertilization procedures. Presumptive zygotes were cultured in groups of 25 in 25-μL droplets of synthetic oviduct fluid supplemented with 5% FCS at 39°C, 5% CO2, and 5%O2 with maximum humidity. In vivo blastocysts were collected from superovulated heifers on Day 7 (estrus = Day 0) by uterine flushing and on Day 13 immediately after slaughter by flushing the dissected reproductive tracts. All embryos were frozen in LN2 and stored at –80°C for mRNA extraction. Quantification of transcripts for RHAMM and CD44 receptors and Has2 and Has3 synthases was performed on groups of ten Day 7 blastocysts (3 groups for in vivo or in vitro) and individual Day 13 embryos (7 embryos in vivo or in vitro) by real-time quantitative RT-PCR. Data on differences in transcript abundance were analyzed by ANOVA. The relative abundance of the Has2 and Has3 synthases was similar between in vivo and in vitro embryos, irrespective of their developmental stage. The quantity of CD44 was significantly higher in in vitro compared with in vivo embryos only on Day 7. However, the quantity of RHAMM receptor was higher on Day 13 in in vitro compared with in vivo embryos. When the comparison was done between developmental stages (Day 7 v. Day 13) for in vivo and in vitro embryos, we found that in vivo-produced Day 7 blastocysts expressed significantly more RHAMM receptor than embryos on Day 13. The reverse pattern of expression was shown for CD44 receptor. For in vitro embryos, the only difference observed was for Has3, which was up-regulated on Day 13 compared with Day 7 embryos. In conclusion, the present study demonstrates, for the first time, developmental changes in the abundance of RHAMM and CD44 receptor mRNA and Has2 and Has3 synthase mRNA in in vivo and in vitro bovine-derived embryos on Day 7 and 13. We believe that our results will provide new insight into the potential role of this intriguing multifunctional molecule in bovine early embryo development.

scholarly journals β-adrenergic Receptor Stimulation Revealed a Novel Regulatory Pathway via Suppressing Histone Deacetylase 3 to Induce Uncoupling Protein 1 Expression in Mice Beige Adipocyte

2018 ◽  
Vol 19 (8) ◽  
pp. 2436 ◽  
Author(s):  
Ana Yuliana ◽  
Huei-Fen Jheng ◽  
Satoko Kawarasaki ◽  
Wataru Nomura ◽  
Haruya Takahashi ◽  
...  
Keyword(s):  
Histone Deacetylase ◽  
Transcriptional Activation ◽  
Adrenergic Receptor ◽  
Uncoupling Protein ◽  
Open Chromatin ◽  
Uncoupling Protein 1 ◽  
Beige Adipocyte ◽  
Ucp1 Expression

Browning of adipose tissue has been prescribed as a potential way to treat obesity, marked by the upregulation of uncoupling protein 1 (Ucp1). Several reports have suggested that histone deacetylase (HDAC) might regulate Ucp1 by remodelling chromatin structure, although the mechanism remains unclear. Herein, we investigate the effect of β-adrenergic receptor (β-AR) activation on the chromatin state of beige adipocyte. β-AR-stimulated Ucp1 expression via cold (in vivo) and isoproterenol (in vitro) resulted in acetylation of histone activation mark H3K27. H3K27 acetylation was also seen within Ucp1 promoter upon isoproterenol addition, favouring open chromatin for Ucp1 transcriptional activation. This result was found to be associated with the downregulation of class I HDAC mRNA, particularly Hdac3 and Hdac8. Further investigation showed that although HDAC8 activity decreased, Ucp1 expression was not altered when HDAC8 was activated or inhibited. In contrast, HDAC3 mRNA and protein levels were simultaneously downregulated upon isoproterenol addition, resulting in reduced recruitment of HDAC3 to the Ucp1 enhancer region, causing an increased H3K27 acetylation for Ucp1 upregulation. The importance of HDAC3 inhibition was confirmed through the enhanced Ucp1 expression when the cells were treated with HDAC3 inhibitor. This study highlights the novel mechanism of HDAC3-regulated Ucp1 expression during β-AR stimulation.

scholarly journals 245RELATIVE ABUNDANCE OF HSP 70 MRNA IN BOVINE EMBRYOS PRODUCED IN VITRO USING DIFFERENT EMBRYO/VOLUME RATIOS IN CULTURE

2004 ◽  
Vol 16 (2) ◽  
pp. 243
Author(s):  
A.T.D. Oliveira ◽  
C. Gebert ◽  
R.F.F. Lopes ◽  
H. Niemann ◽  
J.L. Rodrigues
Keyword(s):  
Relative Abundance ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Hatching Rate ◽  
Bovine Embryos ◽  
Blastocyst Formation ◽  
Cleavage Rate ◽  
Gene Transcripts

In spite of in vitro embryo production systems having been greatly improved over recent years, employing a variety of culture conditions (media, protein sources, gas atmosphere, etc.), we still do not know much about the real necessity of embryos to develop under the same conditions as occur in vivo. These differences between in vivo and in vitro culture at preimplantation embryonic stages can produce deviations in gene expression and in normal fetal development (large offspring syndrome). Heat shock proteins (Hsp) are engaged in cell response to regulatory signals or perturbations in the microenviroment and can be used as a sensitive indicator of stress caused by suboptimal culture conditions (Wrenzycki et al., 2001Hum. Reprod. 16, 893–901). Hsp act as chaperones in facilitating protein folding and assembly and stabilize damaged proteins to prevent aggregation of fragments, thereby allowing repair or degradation. The aim of the present study was to investigate the effects of different embryo/volume ratios on bovine embryo development and the relative abundance of Hsp 70.1 gene transcripts. In this experiment, oocytes were isolated from slaugterhouse ovaries and matured, fertilized and cultured in groups of 5, 10, 20 or 30 per each drop of 100μL. The oocytes were matured in TCM 199 supplemented with 0.4% BSA. After maturation, oocytes were fertilized in TALP medium, using frozen/thawed sperm, selected using a percoll density gradient. The zygotes were cultured to the morula or Day 7 blastocyst stage employing SOF supplemented with 0.4 % BSA. Developmental check points were cleavage rate (Day 3pi), blastocyst formation (Day 8pi) and hatching (Day 11pi). A semi-quantitative RT-PCR assay was used to determine the relative levels of gene transcripts in single embryos at morula (Day 6) and blastocyst (Day 7) stages (Wrenzycki et al., 2001 Biol. Reprod. 65, 309–317). Data of cleavage, blastocyst formation and hatching rates were analyzed using chi-square test. Relative abundance (RA) of Hsp 70.1mRNA were compared in tested groups using ANOVA followed a Tukey test. Differences at P&lt;0.05 were considered significant. Results show that no significative difference in hatching rate per blastocyst produced was detected among the four groups. Cleavage rate and blastocyst formation were significantly higher in groups with 5, 10 and 20 embryos compared with drops containing 30 embryos. Hsp transcripts were detected in morula and blastocyst stages in all groups. In morula stage, no differences were observed in the RA of Hsp 70.1mRNA among groups with 5, 10, 20 and 30 embryos cultured per drop. However, in blastocyst stage, the RA was significantly increased in the group with 20 embryos per drop as compared to the group with 5 embryos. The results show that different embryo/volume ratios in culture influence not only cleavage rate, blastocyst formation and hatching rate, but also expression of Hsp 70.1 gene. Further studies changing other culture conditions and using in vivo-derived bovine embryos will aid in elucidating which culture systems are ideal to produce bovine embryos in vitro. This research was supported by CAPES/DAAD program and CNPq.

scholarly journals 243TEMPORAL DIVERGENCE IN THE PATTERN OF MRNA EXPRESSION IN BOVINE EMBRYOS CULTURED FROM THE ZYGOTE TO BLASTOCYST STAGE IN VITRO OR IN VIVO

2004 ◽  
Vol 16 (2) ◽  
pp. 242
Author(s):  
P. Lonergan ◽  
D. Rizos ◽  
A. Gutierrez-Adan ◽  
P.M. Moreira ◽  
B. Pintado ◽  
...  
Keyword(s):  
Growth Factor ◽  
Relative Abundance ◽  
Transcript Abundance ◽  
Blastocyst Stage ◽  
Insulin Like Growth Factor ◽  
Bovine Embryos ◽  
Interferon Tau ◽  
Cell Stage

The objective of this study was to examine the time during the post-fertilization culture period that gene expression patterns of in vitro cultured bovine embryos diverge from those of their in vivo cultured counterparts. Presumptive bovine zygotes were produced by IVM/IVF of immature oocytes collected from the ovaries of slaughtered animals. At approximately 20h post-insemination (hpi), presumptive zygotes were randomly divided into two culture groups, either in vitro in synthetic oviduct fluid or in vivo, and transferred into the ewe oviduct. Embryos were recovered from both systems at approximately 30hpi (2-cell), two (4-cell), three (8-cell), four (16-cell), five (early morula), six (compact morula) or seven (blastocyst) days pi and snap-frozen for the analysis of transcript abundance using real-time PCR. The transcripts studied were interferon-tau, apoptosis regulator box-a (Bax), connexin 43, sarcosine oxidase, glucose transporter 5, mitochondrial Mn-superoxide dismutase, insulin-like growth factor II, and insulin-like growth factor-I receptor, most of which are known from our previous work to be differentially transcribed in blastocysts derived from culture in vitro or in vivo. Analysis was done on pools of 10 embryos. Data were analyzed using one-way repeated measures ANOVA. The relative abundance of the transcripts studied varied throughout the preimplantation period and was strongly influenced by the culture environment. For example, transcripts for interferon-tau were detected from the 8-cell stage onwards in in vitro-cultured embryos but not until the early morula stage in those cultured in vivo. Levels of this transcript increased significantly at the compact morula and blastocyst stages in both groups but were significantly higher (P&lt;0.05) in in vitro-cultured embryos at both stages. mRNA for Bax was not detected before the 8-cell stage in in vitro cultured embryos and not until the 16-cell stage in in vivo cultured embryos. The abundance of this transcript increased significantly thereafter up to the blastocyst stage in both groups. The level of expression was significantly higher (P&lt;0.05) at all stages of development in in vitro-cultured embryos than those cultured in vivo. The relative abundance of Cx43 transcripts decreased in both in vitro- and in vivo-cultured embryos at the 8- to 16-cell stage. Levels remained low thereafter in the in vitro-cultured embryos but significantly increased in those cultured in vivo. Transcript abundance was significantly higher in in vivo cultured embryos from Day 4 onwards with a ten-fold difference presence at the blastocyst stage. Differences also existed for the other transcripts studied. These data demonstrate that changes in transcript abundance in blastocyst stage embryos are in many cases a consequence of perturbed transcription earlier in development. Depending on the transcript, these differences may be evident in as short as 10h of culture.

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