Characterization of deletions of the HBB locus by array comparative genomic hybridization
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Daniel Sabath
PROVIDER: E-GEOD-68587 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:β-thalassemia major can be caused by homozygous mutations of the HBB gene, most of the cases are inherited from parents who both have β-thalassemia minor. Herein, we show that a mosaic paternal uniparental isodisomy of chromosome 11p14.3-15.5 is associated with β-thalassemia major in a patient with β-thalassemia minor-that evolved to β-thalassemia major. From this case, we suggest that analysis of HBB gene for non-hematopoietic tissues should be performed in late-onset β-thalassemia major patients. Experiment Overall Design: This study is to evaluate the cause of delay-onset β-thalassemia major in our patient. Patients peripheral blood, hair follicle, and oral mucosa, and her parents pripheral blood samples were analyzed.
2008-06-15 | E-GEOD-7847 | biostudies-arrayexpress
Project description:β-thalassemia major can be caused by homozygous mutations of the HBB gene, most of the cases are inherited from parents who both have β-thalassemia minor. Herein, we show that a mosaic paternal uniparental isodisomy of chromosome 11p14.3-15.5 is associated with β-thalassemia major in a patient with β-thalassemia minor-that evolved to β-thalassemia major. From this case, we suggest that analysis of HBB gene for non-hematopoietic tissues should be performed in late-onset β-thalassemia major patients. Keywords: genomic
2007-05-22 | GSE7847 | GEO
Project description:Array-CGH used to characterize HBB deletions causing beta thalassemia and HPFH
2015-05-06 | GSE68587 | GEO
Project description:β-hemoglobinopathies are the most common genetic disorders worldwide. In sickle cell disease (SCD) a single mutation (E6V) in the b-globin (HBB) gene results in dysfunctional hemoglobin protein, while in β-thalassemia, over 300 mutations distributed across the HBB gene are known to reduce the production of β-globin and cause severe anemia. A genetic engineering approach that replaces the whole HBB gene is an ideal strategy to rescue HBB expression for most genotypes but is technically challenging as the insert cannot be homologous to the endogenous gene and codon-optimized, intron-less sequences may not reconstitute adequate HBB levels. Here, we developed a novel approach for a “one-size-fits-all" HBB gene repair strategy using CRISPR-Cas9 which successfully addresses these problems. First, our DNA donor design avoids sequence homology through a diverged HBB coding sequence and second, incorporates heterologous introns from the fetal g-globin gene, further reducing homology to endogenous HBB while mimicking its intron composition. Screening DNA donors with various heterologous globin introns, polyadenylation signals and truncated sequences, identified a heterologous intron DNA donor that highly expresses β-globin and rescued β-globin expression in two in vitro hemoglobinopathy models in hematopoietic stem and progenitor cells (HSPCs). Furthermore, healthy donor HSPCs modified with this HBB gene replacement approach showed successful engraftment in immunodeficient mice at 16 weeks. In summary, we developed a universal HBB gene replacement strategy that results in physiological b-globin production, offering a potential differentiated approach for treating patients with β-thalassemia, SCD or compound heterozygous individuals.
2024-04-01 | GSE229212 | GEO
Project description:Background: The thalassemias are highly diverse at both the molecular and clinical levels. Many of the HBB mutations that result in β-thalassemia are missense mutations in the coding region of the β-globin gene, but a few cause alternative splicing, and interfere with normal processing of the β-globin transcripts. Transcriptome profiling in individuals affected with β-thalassemia, especially in individuals who carry novel mutations in the HBB, may improve our understanding of the heterogeneity and molecular mechanisms of the disease. Methods: Members of a family with a daughter affected with thalassemia intermedia, although her mother was not clinically affected, were examined for physical characteristics, hematological parameters and β-globin gene sequences. We also characterized genome-wide gene expression in the family using RT-qPCR and high-throughput RNA-sequencing mRNA expression profiling of blood. Results: Clinical findings, hematological indices, DNA and RNA sequence analysis of individuals with β-thalassemia, including the description of a novel mutation in the β-globin gene, which introduces a cryptic donor splice site. More than 300 genes are differentially expressed in β-thalassemic blood with many of the DEGs involved in pathways relevant to the clinical management of β-thalassemia. β-thalassemia shows important similarities and differences with sickle cell disease at the transcriptome level. Conclusions: We described the down-regulation of the β-globin gene in β-thalassemia by RNA-sequencing analysis using a sample from an affected individual and her mother, who have a novel mutation in the HBB that creates a cryptic donor splice site. The daughter has a typical β-thalassemia allele as well, and an unexpectedly severe phenotype. The DEGs are enriched in pathways that are directly or indirectly related to β-thalassemia such as hemopoiesis, heme biosynthesis, response to oxidative stress, inflammatory responses, immune responses, control of circadian rhythm, apoptosis, and other cellular activities. We compare our findings with published results of RNA-Sequencing analysis of sickle cell disease (SCD) and erythroblasts from a KLF1-null neonate with hydrops fetalis, and recognize similarities and differences in their transcriptional expression patterns.
2019-03-09 | GSE96060 | GEO
Project description:To investigate the cytogenetic and large-scale chromosomal changes in involuted or non-involuted microGISTs using post-whole genome amplification (WGA) FFPE DNA materials Sixteen patients, total 19 FFPE tumor samples (block storage time 4 months to 9 years), including 16 microGISTs and 3 GISTs larger than 1 cm from the same patients harboring microGISTs. All FFPE tumor samples underwent DNA extraction and WGA (modified degenerate oligonucleotide PCR (DOP) method, provided by Sigma). For each tumor sample, a post-WGA DNA extract from the normal tissue in the same block (or block from the same patient with a block storage time differences less than 2 years) was obtained for tumor sample DNA co-hybridization. Tumor and normal areas of interest were marked and collected from 5- to 10-micron unstained or hematoxylin-stained sections by manual or laser (PixCell IITM, Arcturus Bioscience, CA, USA) microdissection. DNAs were then extracted. WGA was performed using GenomePlex® Tissue Whole Genome Amplification WGA5 kit (Sigma, Saint Louis, MO, USA; http://www.sigmaaldrich.com/) in parallel in accordance with the manufacturer's protocols. At least four independent experiments were concurrently performed per template amplification. Four separate WGA reaction products were pooled for each sample.
2011-12-31 | E-GEOD-28469 | biostudies-arrayexpress
Project description:Development of the human gut microbiota commences at birth with bifidobacteria being among the first colonizers of the sterile newborn gastrointestinal tract. To date the genetic basis of bifidobacterial colonization and persistence remains poorly understood. Transcriptomic analysis of the 2,422,684 bp genome of Bifidobacterium breve UCC2003, a strain isolated from a nursling stool, during colonization of a mouse model revealed the differential expression of a type IVb or so-called Tad pilus-encoding cluster. Mutational analysis coupled with colonization experiments demonstrated that the UCC2003 tad gene cluster is essential for colonization. Immunogold transmission electron microscopy confirmed the presence of the Tad pili at the cell poles of B. breve UCC2003. Conservation of the Tad pilus-encoding locus among sequenced bifidobacterial genomes supports the notion of a ubiquitous and novel host colonization and persistence mechanism for bifidobacteria. Colonisation of BalbC mice: Seven-week-old male, BalbC mice were housed in individually vented cages (Animal care systems, Colorado) under a strict 12 h light cycle. Mice (n = 5 per group) were fed a standard polysaccharide-rich mouse chow diet and water ad libidum. Mice were inoculated by oral gavage (109 cfu of B. breve UCC2003 pPKCM7 or B. breve UCC 2003-tadA pPKCM7 in 100 ul of PBS). Fecal pellets were collected at intervals over 25 days to enumerate bacteria. Twenty five days after inoculation, mice were sacrificed and their intestinal tracts quickly dissected. Aliquots of small intestine, caecum and large intestine were harvested for determination of colony forming units (cfu) (serial dilution plating on RCA agar plates with appropriate antibiotics). Caeca were placed in RNA later for immediate RNA isolation (see below). Microarray procedures: DNA microarrays containing oligonucleotide primers representing each of the 1926 annotated genes of B. breve UCC2003 were obtained from Agilent Technologies (Palo Alto, CA). An overnight culture of B. breve was used to inoculate (1 % inoculum) 50 ml of MRS broth. Cells were incubated at 37°C until an optical density at 600 nm (OD600) of 0.5 was reached. Cells were then harvested by centrifugation at 8,000 x g for 1 min at room temperature and immediately frozen at -80oC. Methods for cell disruption, RNA isolation, RNA quality control were performed as described previously (van Hijum et al., 2005). Caeca, isolated from mice, were placed in RNAlater (Ambion) and immediately processed. Bacterial mRNA was extracted from caecal RNA preparations using the MicrobEnrich and MicrobExpress kits (Ambion) according to the manufacturer’s instructions. cDNA from bacterial mRNA was synthesized using the cDNA synthesis and labelling kit DSK-001 (Kreatech) according to the manufacturer’s instructions. 1 μl of each cDNA solution (10 ng) was amplified in triplicate using the GenomiPhi™ V2 DNA Amplification Kit (Amersham Biosciences) according to the manufacturer's protocol and labelled with Cy3 or Cy5 using Cy3-ULS and Cy5-ULS from the cDNA synthesis and labelling kit DSK-001 (Kreatech). Labelled and amplified cDNA was hybridized using the Agilent Gene Expression hybridization kit (part number 5188-5242) as described in the Agilent Two-Color Microarray-Based Gene Expression Analysis (v4.0) manual (publication number G4140-90050). Following hybridization, microarrays were washed as described in the manuals and scanned using Agilent's DNA microarray scanner G2565A. The scanning results were converted to data files with Agilent's Feature Extraction software (version 9.5). DNA microarray data was processed as previously described (Zomer et al. 2009). Differential expression tests were performed using a t test. A gene was considered differentially expressed between a test strain and control when an expression ratio of >5 or <0.2 relative to the result for the control was obtained with a corresponding P value that was <0.0001. DNA microarrays containing oligonucleotide primers representing each of the 1926 annotated genes of B. breve UCC2003 were obtained from Agilent Technologies (Palo Alto, CA). An overnight culture of B. breve was used to inoculate (1 % inoculum) 50 ml of MRS broth. Cells were incubated at 37°C until an optical density at 600 nm (OD600) of 0.5 was reached. Cells were then harvested by centrifugation at 8,000 x g for 1 min at room temperature and immediately frozen at -80oC. 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2011-07-01 | E-GEOD-27491 | biostudies-arrayexpress
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2014-10-01 | E-GEOD-60376 | biostudies-arrayexpress
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