HUN-REN BRC CF.

Magyar

Laboratory of Functional Genomics

About us

Since 2000, the Functional Genomics Laboratory at the Szeged Biological Centre has been offering multiple platforms for the individual or systematic analysis of gene expression, nucleic acids (RNA, DNA, microRNA, cell-free cfRNA circulating in blood) and quantitative analysis of DNA methylation. The laboratory carries out its own research, partly in the framework of scientific collaborations, and partly as a service provider. We have several collaborating partners from both national and international institutions.
Our main developments include nucleic acid pattern and signal amplification techniques from sparse biological samples, single-cell RNA sequencing methods, digital nucleic acid detection technologies, gene expression bioinformatics analyses.

Technology

Expertise and Equipment

1. Real-time PCR (qPCR) technology

The real-time PCR technique allows the absolute (but requires a calibration curve) and relative quantification of nucleic acids (DNA, RNA). It can be used for gene expression studies, copy number determination, DNA microarray, RNAseq, validation of results obtained from next-generation sequencing (NGS) experiments, and detection of polymorphisms, SNPs. The method is based on the PCR technique, after each synthesis step, the machine detects the fluorescence intensity of the fluorescent dye bound to the double-stranded DNA present (SybrGreen method) and the fluorescence intensity of the binding candidate from the hybridization assay (TaqMan assay). This fluorescence intensity is proportional to the amount of DNA present in the reaction mixture, making this method suitable for the determination of starting DNA amounts. Our laboratory is equipped with a Corbett Research RotorGene 3000 real-time PCR machine and QuantStudio 3 (Applied Biosystems). RotorGene allows the analysis of 72 samples at a time, but the QuantStudio3 runs with 96 well plates. One of the most important steps in successful quantification is careful primer and assay design and proper interpretation of the data. On a collaborative basis we undertake the design and execution of all experimental steps from primer design to data evaluation.

2. Digital PCR (dPCR) technology

Digital PCR is a PCR-based technology that allows the absolute quantification of very small amounts of nucleic acids without a calibration curve, in contrast to the relative quantification of qPCR, and with high accuracy. By nanolitre dispersion of the reaction mixture, it is possible to obtain one or zero template DNA per reaction unit, so that after appropriate Poisson correction and statistics, the number of molecules in the reaction mixture can be determined. The method is highly sensitive and therefore suitable for single cell genomics, copy number determination, detection of rare sequences, mutations, alleles in basic and clinical research, biomarker and diagnostic studies (cell-free nucleic acids circulating in blood, cfRNA, miRNA). In our laboratory we use the BioRad QX200 Droplet Digital System with all necessary hardware and software background under optimized conditions.

3. RNAscope in situ hibridization (ACDBio)

RNAscope technology is a revolutionary in situ hybridization method for the detection of specific RNA molecules within intact cells. It is characterized by outstanding specificity and sensitivity as well as quantifiability due to its unique assay design. The lower Z-shaped probe section (18-20bp) is specific to the target sequence, which is linked to the upper (14bp) tail sequence by a connecting sequence fragment. Two closely related Z probes together form a 28bp long sequence segment, to which fluorescent dye is attached via a signal multiplier system. The outstanding specificity is due to the fact that only two adjacent hybridizing probes can give a fluorescent signal. The method works on both fresh-frozen and fixed-frozen samples, as well as on samples embedded in paraffin. It is possible to use up to 12 probes simultaneously, so that 12 gene expression patterns can be detected on a single section at a given time. This method is an excellent complement to other quantitative methods for validation. In our laboratory we use the RNAscope system from ACDBio.

Our best

Services

Gene expression assays using focused panels with real-time quantitative PCR recapture or single-cell digital droplet PCR

Our group has qPCR (Corbett Research) and digital droplet PCR (BioRad) available to track the gene expression changes that the user wants to investigate. The service may include primer design or Taqman assay selection, cell handling, RNA isolation, cDNA transcription and qPCR reaction assembly and run. Our service can include interpretation of the results, plotting of differential gene expression patterns, pathway analysis (e.g. KEGG). Single cell-based assays can highlight the heterogeneity of the cells in the sample. Single cell digital qPCR can be used to track the expression of 4 genes per cell.
https://pubmed.ncbi.nlm.nih.gov/27487831/
https://pubmed.ncbi.nlm.nih.gov/30637442/
https://www.nature.com/articles/s41598-019-48888-6

Articles

Key publications

  • 1. Köhler, Zoltán Márton ; Trencsényi, György ; Juhász, László ; Zvara, Ágnes ; Szabó, P Judit ; Dux, László ; Puskás, G László ; Rovó, László ; Keller-Pintér, Anikó, Tilorone increases glucose uptake in vivo and in skeletal muscle cells by enhancing Akt2/AS160 signaling and glucose transporter levels, JOURNAL OF CELLULAR PHYSIOLOGY 238 : 5 pp. 1080-1094. , 15 p. (2023)
  • 2. Shukla, R.D. ; Zvara, Á. ; Avramucz, Á. ; Biketova, A.Yu. ; Nyerges, A. ; Puskás, L.G. ; Fehér, T., inPOSE: A Flexible Toolbox for Chromosomal Cloning and Amplification of Bacterial Transgenes, MICROORGANISMS 10 : 2 Paper: 236 (2022
  • 3. Tóth, Melinda E. ✉ ; Sárközy, Márta* ✉ ; Szűcs, Gergő ; Dukay, Brigitta ; Hajdu, Petra ; Zvara, Ágnes ; Puskás, László G. ; Szebeni, Gábor J. ; Ruppert, Zsófia ; Csonka, Csaba et al. Exercise training worsens cardiac performance in males but does not change ejection fraction and improves hypertrophy in females in a mouse model of metabolic syndrome BIOLOGY OF SEX DIFFERENCES 13 : 1 Paper: 5 , 20 p. (2022)
  • 4. Váczi, S. ; Barna, L. ; Harazin, A. ; Mészáros, M. ; Porkoláb, G. ; Zvara, Á. ; Ónody, R. ; Földesi, I. ; Veszelka, S. ; Penke, B. et al. S1R agonist modulates rat platelet eicosanoid synthesis and aggregation, PLATELETS 33 : 5 pp. 709-718. , 10 p. (2022)
  • 5. Koos K, Oláh G, Balassa T, Mihut N, Rózsa M, Ozsvár A, Tasnadi E, Barzó P, Faragó N, Puskás L, Molnár G, Molnár J, Tamás G, Horvath P. Automatic deep learning-driven label-free image-guided patch clamp system. Nat Commun. 2021 Feb 10;12(1):936. doi: 10.1038/s41467-021-21291-4.
  • 6. Balogh A, Reiniger L, Hetey S, Kiraly P, Toth E, Karaszi K, Juhasz K, Gelencser Z, Zvara A, Szilagyi A, Puskas LG, Matko J, Papp Z, Kovalszky I, Juhasz C, Than NG. Decreased Expression of ZNF554 in Gliomas is Associated with the Activation of Tumor Pathways and Shorter Patient Survival. Int J Mol Sci. 2020 Aug 11;21(16):5762. doi: 10.3390/ijms21165762. PMID: 32796700; PMCID: PMC7461028.
  • 7. Barna L, Walter FR, Harazin A, Bocsik A, Kincses A, Tubak V, Jósvay K, Zvara Á, Campos-Bedolla P, Deli MA. Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage. Fluids Barriers CNS. 2020 Feb 10;17(1):5. doi: 10.1186/s12987-019-0166-1. PMID: 32036791; PMCID: PMC7008534.
  • 8. Gyukity-Sebestyén E, Harmati M, Dobra G, Németh IB, Mihály J, Zvara Á, Hunyadi-Gulyás É, Katona R, Nagy I, Horváth P, Bálind Á, Szkalisity Á, Kovács M, Pankotai T, Borsos B, Erdélyi M, Szegletes Z, Veréb ZJ, Buzás EI, Kemény L, Bíró T, Buzás K. Melanoma-Derived Exosomes Induce PD-1 Overexpression and Tumor Progression via Mesenchymal Stem Cell Oncogenic Reprogramming. Front Immunol. 2019 Oct 18;10:2459. doi: 10.3389/fimmu.2019.02459. PMID: 31681332; PMCID: PMC6813737.
  • 9. Bencsik P, Kiss K, Ágg B, Baán JA, Ágoston G, Varga A, Gömöri K, Mendler L, Faragó N, Zvara Á, Sántha P, Puskás LG, Jancsó G, Ferdinandy P. Sensory Neuropathy Affects Cardiac miRNA Expression Network Targeting IGF-1, SLC2a-12, EIF-4e, and ULK-2 mRNAs. Int J Mol Sci. 2019 Feb 25;20(4):991. doi: 10.3390/ijms20040991. PMID: 30823517; PMCID: PMC6412859.
  • 10. Fenteany, Gabriel* ; Gaur, Paras* ; Hegedűs, Lili ; Dudás, Kata ; Kiss, Ernő ; Wéber,Edit ; Hackler, László ; Martinek, Tamás ; Puskás, László G. ; Haracska, Lajos Multilevel structure–activity pro􀅣ling reveals multiple green tea compound familiesthat each modulate ubiquitin-activating enzyme and ubiquitination by a distinctmechanism, SCIENTIFIC REPORTS 9 : 1 Paper: 12801 , 16 p. (2019)
  • 11. Sárközy M, Gáspár R, Zvara Á, Kiscsatári L, Varga Z, Kővári B, Kovács MG, Szűcs G, Fábián G, Diószegi P, Cserni G, Puskás LG, Thum T, Kahán Z, Csont T, Bátkai S. Selective Heart Irradiation Induces Cardiac Overexpression of the Pro- hypertrophic miR-212. Front Oncol. 2019 Jul 16;9:598. doi: 10.3389/fonc.2019.00598. PMID: 31380269; PMCID: PMC6646706.
  • 12. Sárközy M, Gáspár R, Zvara Á, Siska A, Kővári B, Szűcs G, Márványkövi F, Kovács MG, Diószegi P, Bodai L, Zsindely N, Pipicz M, Gömöri K, Kiss K, Bencsik P, Cserni G, Puskás LG, Földesi I, Thum T, Bátkai S, Csont T. Chronic kidney disease induces left ventricular overexpression of the pro-hypertrophic microRNA-212. Sci Rep. 2019 Feb 4;9(1):1302. doi: 10.1038/s41598-018-37690-5. PMID: 30718600; PMCID: PMC6362219.
  • 13. Csajbók ÉA, Kocsis ÁK, Faragó N, Furdan S, Kovács B, Lovas S, Molnár G, Likó I, Zvara Á, Puskás LG, Patócs A, Tamás G. Expression of GLP-1 receptors in insulin-containing interneurons of rat cerebral cortex. Diabetologia. 2019 Apr;62(4):717-725. doi: 10.1007/s00125-018-4803-z. Epub 2019 Jan 12. PMID: 30637442. Intelligent image-based in situ single-cell isolation.
  • 14. Brasko C, Smith K, Molnar C, Farago N, Hegedus L, Balind A, Balassa T, Szkalisity A, Sukosd F, Kocsis K, Balint B, Paavolainen L, Enyedi MZ, Nagy I, Puskas LG, Haracska L, Tamas G, Horvath P., Nat Commun. 2018 Jan 15;9(1):226. doi: 10.1038/s41467-017-02628-4.

Team

Check our Team

László PUSKÁS

scientific adviser

Gábor SZEBENI

senior research associate

Ágnes ZVARA

senior research associate

Fanni BALOGH

research associate

Nóra Katalin FARAGÓ

research associate

Beáta KARI

research associate

Nikolett GÉMES

research associate

Patrícia NEUPERGER

research associate

Edit, KOTOGÁNY

junior research associate

Beáta MANDÁKNÉ MILITÁR

laboratory assistant/administrator expert

Cynthia SZABÓ

laboratory assistant/administrator expert