Medizinstudierende als Helfende in der Pandemie

Die Anaesthesiologie - Die COVID-19-Pandemie hat das deutsche Gesundheitssystem vor enorme Herausforderungen gestellt und den Bedarf an Strategien zu Rekrutierung, Schulung und Einsatzplanung von...     

Establishing signs for acute and healing phases of distal tibial classic metaphyseal lesions

Pediatric Radiology - Stages of healing for classic metaphyseal lesions (CMLs) are not well established. Follow-up skeletal surveys provide an opportunity to evaluate signs of healing CMLs. To...     

Development of an effective and rapid qPCR for identifying human ChREBPα/β isoforms in hepatic and adipose tissues

Most quantitative real-time PCR (qPCR) detection methods use two types of chemistries to measure the expression levels of ChREBP isoforms, hydrolysis probes for ChREBPα and SYBR Green for ChREBPβ. Hydrolysis probes are not available to determine the ChREBPβ isoform. The aim of this study was to develop a qPCR assay based only on hydrolysis probes for both ChREBP isoforms. Liver and adipose tissue biopsies from patients undergoing elective cholecystectomy surgery were used to perform qPCR. To validate this assay, the results were compared with sequencing and High Resolution Melting (HRM) PCR assays. Direct sequencing was used to determine the sequence showing site where ChREBPβ presents its specific splicing (1?b exon/2 exon) in order to design the primers and the probe. We developed a qPCR assay to determine the ChREBP isoforms expression based on hydrolysis probes. It assays showed good efficiency (95.50%, on average), high reproducibility, and a strong linear correlation (R² ≥ 0.99) for tissues tested. HRM analysis confirmed the specificity of the primers and the result of this assay matched (100%) with the outcomes obtained by sequencing and qPCR. Also, we obtained the ChREBPβ sequence showing exon 1b spliced to exon 2, bypassing exon 1a, and retaining the remainder of the ChREBPα exons. Based on the use of hydrolysis probes, our method can efficiently identify the expression of both ChREBP isoforms. Thus, the comparability of the qPCR results using a single chemistry (hydrolysis probes) to discriminate between both ChREBP isoforms was possible.