Project description:Velo-cardio-facial syndrome/DiGeorge syndrome/22q11.2 deletion syndrome (22q11DS) patients have a submucous cleft palate, velo-pharyngeal insufficiency associated with hypernasal speech, facial muscle hypotonia and feeding difficulties. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on 22q11.2, results in a cleft palate and a reduction or loss of branchiomeric muscles. To identify genes downstream of Tbx1 for myogenesis, gene profiling was performed on mandibular arches (MdPA1) from Tbx1+/+ and Tbx1-/- mouse embryos. To obtain enough RNA for microarray hybridization experiments, dissected mandibular arches from three Tbx1+/+ and three Tbx1-/- E9.5 embryos were pooled according to genotype, with three microarrays performed in total per genotype. Affymetrix Mouse Gene ST 1.0 arrays (Affymetrix) were used. Hybridization, washing, staining and scanning were performed in the Genomics Core at Einstein (http://www.einstein.yu.edu/genetics/CoreFacilities.aspx?id=23934) according to the Affymetrix manual.
Project description:Velo-cardio-facial syndrome/DiGeorge syndrome/22q11.2 deletion syndrome (22q11DS) patients have a submucous cleft palate, velo-pharyngeal insufficiency associated with hypernasal speech, facial muscle hypotonia and feeding difficulties. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on 22q11.2, results in a cleft palate and a reduction or loss of branchiomeric muscles. To identify genes downstream of Tbx1 for myogenesis, gene profiling was performed on mandibular arches (MdPA1) from Tbx1+/+ and Tbx1-/- mouse embryos.
Project description:Velo-cardio-facial syndrome/DiGeorge syndrome/22q11.2 deletion syndrome (22q11DS) patients have a submucous cleft palate, velo-pharyngeal insufficiency associated with hypernasal speech, facial muscle hypotonia and feeding difficulties. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on 22q11.2, results in a cleft palate and a reduction or loss of branchiomeric muscles. To identify genes downstream of Tbx1 for myogenesis, gene profiling was performed on mandibular arches (MdPA1) from Tbx1+/+ and Tbx1-/- mouse embryos.
Project description:Velo-cardio-facial syndrome/DiGeorge syndrome/22q11.2 deletion syndrome (22q11DS) patients have a submucous cleft palate, velo-pharyngeal insufficiency associated with hypernasal speech, facial muscle hypotonia and feeding difficulties. Inactivation of both alleles of mouse Tbx1, encoding a T-box transcription factor, deleted on 22q11.2, results in a cleft palate and a reduction or loss of branchiomeric muscles. To identify genes downstream of Tbx1 for myogenesis, gene profiling was performed on mandibular arches (MdPA1) from Tbx1+/+ and Tbx1-/- mouse embryos. To obtain enough RNA for microarray hybridization experiments, dissected mandibular arches from three Tbx1+/+ and three Tbx1-/- E10.5 embryos were pooled according to genotype, with three microarrays performed in total per genotype. The tissue was homogenized in Buffer RLT (QIAGEN). Total RNA was isolated with the RNeasy Micro Kit according to the manufacturer’s protocol. Quality and quantity of total RNA was determined using an Agilent 2100 Bioanalyzer (Agilent) and an ND-1000 Spectrophotometer (NanoDrop), respectively. Biotinylated single-stranded cDNA targets were amplified from 100 nanograms (ng) starting total RNA using the Ovation RNA Amplification System V2 and FL- Ovation cDNA Biotin Module V2 (NuGEN). A total of 3.75 ?g of cDNA from the last step was hybridized to the GeneChip Test3 array (Affymetrix) to test the quality of the labeled target. Nucleic acid samples that passed quality control were then hybridized to the GeneChip Mouse Genome 430 2.0 Arrays (Affymetrix). Hybridization, washing, staining and scanning were performed in the Genomics Core at Einstein (http://www.einstein.yu.edu/genetics/CoreFacilities.aspx?id=23934) according to the Affymetrix manual.
Project description:We present a gene expression atlas of early mouse craniofacial development. Laser capture microdissection (LCM) was used to isolate cells from the principal critical micro-regions, whose development, differentiation and signaling interactions are responsible for the construction of the mammalian face We examined the facial mesenchyme and adjacent neuroepithelium at E8.5, at E9.5 we obtain cells from the facial mesenchyme, olfactory placode/epidermal ectoderm, underlying neuroepithileium, and emerging mandibular and maxillary arches. AT E10.5 we sampled the medial and lateral prominences, olfactory pit, multiple regions of the underlying neuroepithelium the mandibular and maxillary arches, and Rathke's pouch. Mouse emrbyos were harvested at developmental stage E8.5 , E9.5, and E10.5 and cells were captured from microregions responsible for the construction of the mammalian face. RNA was extracted, labelled, and quantified using the Mouse ST-l microarray.
Project description:This study was aimed at identifying Tbx1 dosage-dependent genes in vivo, so we performed a transcriptome analysis of Tbx1 mutants with nine different genotypes corresponding to different Tbx1 mRNA dosages. RNA isolated from whole E9.5 embryos with 9 different genotypes was hibridized to Affymetrix GeneChip Mouse Genome 430 2.0 arrays. For each genotype, we used two embryos (biological replicates), each hybridized to one array, thus we analyzed a total of 18 arrays. Tbx1 gene dosage