Methodological focus
The methodological focus of the laboratory is the epigenetic characterization of DNA samples. Over the last 10 years, bisulphite sequencing (according to Sanger) has been used to analyze methylation patterns in the promoter region of certain genes. Currently, nanopore sequencing has also been established in our laboratory in order to be able to analyze larger DNA regions using Cas9-mediated enrichment. This method also makes it possible to sequence the entire genome. If deviating methylation patterns are detected in a clinical picture, the functional relevance or the effect at the expression level is checked using real-time PCR, Western blot, ELISA and other gene expression analyses and correlated with the psychometric data. In this way, correlations between the regulation of relevant genes and the expression of psychologically abnormal behavioral patterns can be demonstrated.
The Molecular Neuroscience Laboratory is used by many of the Department's research groups. In particular, the research group "Molecular Psychiatry" research group.
The equipment of the laboratory
The laboratory is equipped with two Biomek Nxp robots from Beckman-Coulter . Thanks to the versatile programmability of these auxiliary robots, the following processes can be carried out:
- Purification of DNA and RNA from sample material and
- Preparation and post-processing of PCRs and sequencing PCRs,
- largely all pipetting steps, which can be performed identically for a number of samples.
In all these processes, automation achieves more consistent and stable results compared to manual processing and higher throughput.
We have two sequencing methods available for DNA analysis:
Nanopore sequencing has been established in our laboratory for analyzing base sequences and methylation patterns of larger DNA regions using Cas9-mediated enrichment or the entire genome without further chemical conversion and PCR amplification.
Further DNA analyses (bisulfite sequencing, SNP, VNTR, LPR) using Sanger sequencing are carried out on an ABI 3500xL sequencer from Applied Biosystems.
For the amplification of nucleic acids, we have four thermal cyclers and a real-time cycler from BioRad at our disposal. An electrophoresis system (chambers and PowerPac™ Basic power supply) and GelDoc Imaging System from BioRad are used to visualize the PCR products.
DNA/RNA/protein quantification is carried out using a DS-11 series spectrophotometer/fluorometer from Denovix.
The Bioanalyzer 2100 from Agilent is used to determine the size and quality of DNA and RNA more precisely.
For the concentration of low-concentration (DNA) samples, our laboratory uses the Savant DNA 120 Speedvac Concentrator from Thermo Scientific.
Protein detection and protein quantification are carried out using protein gel and blot chambers, the VersaDoc Imaging System from BioRad and an ELISA device from Molecular Devices including a plate washer from BioTek.
For the cultivation of cell lines, primary cells and iPS cells, we have a fully equipped S1 laboratory with sterile workbenches, incubators, centrifuge, microscope and live cell imager.
For the cloning and expression of recombinant proteins as well as restriction, ligation and transformation, two rooms in our laboratory are equipped for safety level 1 genetic engineering work. CRISPR/Cas9 technology, gene expression and promoter activity studies using the GloMax Multi Detection System from Promega and genotyping are also used here. Incubators from Memmert, a temperature-controlled shaker from Edmund Bühler GmbH including an incubation hood for cultivating (recombinant) microorganisms and an electroporator for transfection are available for these methods.
The laboratory has five Sanyo ultra-low temperature freezers for storing sample material. The sample material can be managed sustainably using the MySamples software by storing it in Landgraf drawer inserts specially designed for the low temperature range.