Project description:The Del-Mar 14K chip was used to interrogate differential expression of transcripts in the white isthmus (WI) compared with the adjacent magnum (Mg) and uterine (Ut) segments of the hen oviduct. Differential expression of genes common to both comparisons (WI/Mg and WI/Ut) was detected for 204 annotated proteins. Of these, 58 genes were overexpressed in both WI/Mg and WI/Ut, and are therefore considered to be the most interesting candidates for WI - specific functions. Additionally, general analysis revealed 135 clones hybridizing to overexpressed transcripts (WI/Mg + WI/Ut), and corresponding to 102 NCBI annotatated non-redundant Gallus gallus gene ID~s. This combined analysis revealed that structural proteins highly over-expressed in white isthmus were collagen X (COL10A1), Fibrillin (FBN1) and Cysteine Rich Eggshell Membrane Protein (CREMP). In addition, genes encoding collagen-processing enzymes were over-expressed, as were proteins known to regulate disulfide cross-linking, suggesting that coordinated upregulation of gene networks in the white isthmus is associated with eggshell membrane fibre formation. IPA interactome analysis reinforces the key role of the estrogen receptor and SMAD3 in mediating gene regulation during eggshell membrane synthesis. These results will assist with development of selection strategies to improve eggshell quality and food safety of the table egg. Keywords: Laying hen, eggshell, oviduct, Isthmus expression, cDNA microarray, indirect cDNA labelling, Alexa Fluor dyes Keywords: Expression profiling by array
Project description:The Del-Mar 14K chip was used to interrogate differential expression of transcripts in the white isthmus (WI) compared with the adjacent magnum (Mg) and uterine (Ut) segments of the hen oviduct. Differential expression of genes common to both comparisons (WI/Mg and WI/Ut) was detected for 204 annotated proteins. Of these, 58 genes were overexpressed in both WI/Mg and WI/Ut, and are therefore considered to be the most interesting candidates for WI - specific functions. Additionally, general analysis revealed 135 clones hybridizing to overexpressed transcripts (WI/Mg + WI/Ut), and corresponding to 102 NCBI annotatated non-redundant Gallus gallus gene ID~s. This combined analysis revealed that structural proteins highly over-expressed in white isthmus were collagen X (COL10A1), Fibrillin (FBN1) and Cysteine Rich Eggshell Membrane Protein (CREMP). In addition, genes encoding collagen-processing enzymes were over-expressed, as were proteins known to regulate disulfide cross-linking, suggesting that coordinated upregulation of gene networks in the white isthmus is associated with eggshell membrane fibre formation. IPA interactome analysis reinforces the key role of the estrogen receptor and SMAD3 in mediating gene regulation during eggshell membrane synthesis. These results will assist with development of selection strategies to improve eggshell quality and food safety of the table egg. Keywords: Laying hen, eggshell, oviduct, Isthmus expression, cDNA microarray, indirect cDNA labelling, Alexa Fluor dyes Keywords: Expression profiling by array A balanced block hybridization design (Dye switch) was used where half of the samples were labelled with AlexaM-BM-. 555 fluorescent dye and the other half with AlexaM-BM-. 647. A total of 16 microarray slides were used for hybridization to 32 samples that correspond to four tissue contrast (White isthmus versus magnum and uterus versus white isthmus).
Project description:The aim of this study was to assess the impact of oocyte competence on subsequent fertility. Based on knowledge already accessible in mammals and on bioinformatics tools including the chicken genome sequence, we focused on the expression of genes involved in the processes of fertilization and of early embryo development. A differential kinetic study is performed on INRA lines selected on the basis of their fertility potential in purpose of hopefully access gene markers of fertility performance. We use 4 different hen lines:; - one line of laying hens with 3 different samples: the just ovulated oocyte, the oocyte collected 24 hours before ovulation (F1 stage), and granulosa cells collected at the F1 stage. We could compare different tissue and developmental stages. - one line of hen with rapid growth speed; - two lines of laying hens; For the 3 last lines we used animals with different fertility levels. We collected the oocyte of the largest follicle before ovulation (F1). The aim of the study is to identify genes involved in fertility or early embryo mortality. Experiment Overall Design: 6 arrays - Gallus gallus
Project description:The aim of this study was to assess the impact of oocyte competence on subsequent fertility. Based on knowledge already accessible in mammals and on bioinformatics tools including the chicken genome sequence, we focused on the expression of genes involved in the processes of fertilization and of early embryo development. A differential kinetic study is performed on INRA lines selected on the basis of their fertility potential in purpose of hopefully access gene markers of fertility performance. We use 4 different hen lines: - one line of laying hens with 3 different samples: the just ovulated oocyte, the oocyte collected 24 hours before ovulation (F1 stage), and granulosa cells collected at the F1 stage. We could compare different tissue and developmental stages. - one line of hen with rapid growth speed - two lines of laying hens For the 3 last lines we used animals with different fertility levels. We collected the oocyte of the largest follicle before ovulation (F1). The aim of the study is to identify genes involved in fertility or early embryo mortality. Keywords: normal vs disease comparison
Project description:Current understanding of oocyte quality and developmental potential maintains that stochastic or epigenetic processes modulate the action of determinants that are laid in the oocyte during oogenesis. These determinants are considered to be rather conserved across mice, leading different strains of mice to be used interchangeably. We challenged this assumption and studied the relationship between oocyte composition and developmental quality in four inbred strains of mice, namely 129Sv, C57Bl/6, C3H/HeN and DBA/2J. These oocytes showed large variability developmental competence and embryo quality irrespective of the developmental stimulus (fertilization, somatic cell nuclear transfer, parthenogenesis). To unravel the molecular basis of the observed phenotypes we applied state-of-the-art proteomics (SILAC LC-MS/MS) combined with transcriptomics (RNA deep sequencing). We quantified 1839 proteins and 20413 transcripts simultaneously in oocytes of all four strains. The proteome and the transcriptome had little correlation with each other, highlighting the importance of proteomic quantifications in embryology. We found that proteins that were most variably expressed between oocytes from different strains mainly relate to oocyte biology, ribosome and RNA biogenesis as well as embryo differentiation and chromatin remodelling. Thus, different strains of mice should not be used interchangeably in biology when tackling questions about oogenesis and early development.