Methylation profile analysis of Of DNA-HALO structure by synchrotron-based FTIR spectroscopy
2023
Authors:
Sarić, AnaRajić, Jovana
Tolić, Anja
Đorđević, Marija
Grdović, Nevena
Uskoković, Aleksandra
Dinić, Svetlana
Arambašić Jovanović, Jelena
Mihailović, Mirjana
Vidaković, Melita
Dučić, Tanja
Contributors
Dobrijević, ZoranaDocument Type:
Conference object (Published version)
,
© 2023 by the Institute of Molecular Genetics and Genetic Engineering, University of Belgrade
Metadata
Show full item recordAbstract:
Introduction: DNA methylation is a major regulator of transcriptional activity and alongside other epigenetic modifications it introduces specific level of chromatin complexity. Fourier transform infrared (FTIR) spectroscopy is a rapid, non-destructive, and label-free technique for identifying subtle changes in all bio-macromolecules, and it has been used as a method of choice for studying DNA conformation. The present study was designed to explore the use of synchrotron-based FTIR spectroscopy to monitor the subtle changes on molecular level regarding the DNA methylation status of cytosine in the whole genome.
Methods: For FTIR-based DNA methylation analysis in situ, DNA-HALO samples were prepared using slightly modified methodology for nuclear HALO preparations where DNA-HALOs are liberated of any protein residues but preserve higher order chromatin structure.
Results: Using FTIR spectroscopy we analysed and compared methylation profiles of isolated genomic DNA and DNA-HALO samples. DNA-HALO structure shows more distinct peaks in fingerprint region of spectra. DNA-HALO structure is more accurate for detecting bonds in unmemthylated cytosine as specific infrared peaks are defined as vibrations of bonds in unmethylated cytosine at 1151 cm-1 and 1357 cm-1. The ratio of integrated area under the peak 1151 cm-1 over integrated area under the peak that represents PO2-backbone vibrations can be used to assess level of unmethylated cytosine and thus methylation rate in the DNA-HALO samples.
Conclusion: This study demonstrates potential of FTIR spectroscopy to detect DNA methylation in DNA-HALO samples more precisely compared to classical DNA extraction procedure that yield unstructured whole genomic DNA.
Keywords:
HALO structure; FTIR spectroscopy; DNA methylation; Epigenetic marks; ChromatinFunding / projects:
- ALBA In-house grant: “Towards FTIR spectroscopy of single living cells” PI T. Ducic, 2018093034
- COST Action CA1406
- COST Action CA16119
- Ministry of Science, Technological Development and Innovation of the Republic of Serbia, institutional funding - 200007 (University of Belgrade, Institute for Biological Research 'Siniša Stanković') (RS-MESTD-inst-2020-200007)
In:
- Dobrijević Z, editor. Abstract Book: CoMBoS2 - the Second Congress of Molecular Biologists of Serbia; 2023 Oct 6-8; Belgrade, Serbia. Belgrade: Institute of Molecular Genetics and Genetic Engineering, University of Belgrade ; 2023. p. 68. (Trends in Molecular Biology; Special Issue).