Project description:In polygenic disorders we do not know exactly, how many genes are involved in the pathomechanism, but the analysis of fetal gene expression can get us closer to the solution. In our study we were searching for the genetic background of the polygenic neural tube defect, which is the second most common birth defect in the world (1 in 1000 live births). Our data revealed novel candidate genes, like SLAP, LST1 and BENE, which can play an important role in the pathogenesis of neural tube defects. We created a data warehouse from the results, suitable for further analysis. This study also demonstrates that a routinely collected amount of amniotic fluid (as small as 6 mL) is enough to successfully hybridize isolated RNA to expression arrays, making the ability to use the technique from normally collected amniotic fluid samples. Keywords: Prenatal gene expression signature
Project description:In polygenic disorders we do not know exactly, how many genes are involved in the pathomechanism, but the analysis of fetal gene expression can get us closer to the solution. In our study we were searching for the genetic background of the polygenic neural tube defect, which is the second most common birth defect in the world (1 in 1000 live births). Our data revealed novel candidate genes, like SLAP, LST1 and BENE, which can play an important role in the pathogenesis of neural tube defects. We created a data warehouse from the results, suitable for further analysis. This study also demonstrates that a routinely collected amount of amniotic fluid (as small as 6 mL) is enough to successfully hybridize isolated RNA to expression arrays, making the ability to use the technique from normally collected amniotic fluid samples. Experiment Overall Design: We report an in vivo study in living human fetuses using oligonucleotide microarray analysis of fetal mRNA isolated from amniocytes, which were collected by amniocentesis. RNA was successfully isolated, amplified, labeled, and hybridized to whole-genome Affymetrix transcript arrays.
Project description:We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3’ and/or 5’ end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5’ differences and in support of this we report that a 5’ isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5’ isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes
Project description:Single-cell mRNA sequencing was overlaid with single-cell ATAC sequencing of the mouse E9.5 cranial region for both wildtype and miR-302 knockout embryos, an established example of a neural tube closure defect. The goals of this study were to compare changes in gene expression and chromatin accessibility upon loss of a stem cell miRNA and neural tube closure defect.