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Determination of the ribonucleotide content of mtDNA using alkaline gels
Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.ORCID-id: 0000-0001-5704-3093
Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.ORCID-id: 0000-0002-8607-7564
2023 (engelsk)Inngår i: Mitochondrial DNA: methods and protocols / [ed] Thomas J. Nicholls; Jay P. Uhler; Maria Falkenberg, New York: Humana Press, 2023, Vol. 2615, s. 293-314Kapittel i bok, del av antologi (Fagfellevurdert)
Abstract [en]

Impaired mitochondrial DNA (mtDNA) maintenance, due to, e.g., defects in the replication machinery or an insufficient dNTP supply, underlies a number of mitochondrial disorders. The normal process of mtDNA replication leads to the incorporation of multiple single ribonucleotides (rNMPs) per mtDNA molecule. Given that embedded rNMPs alter the stability and properties of the DNA, they may have consequences for mtDNA maintenance and thereby for mitochondrial disease. They also serve as a readout of the intramitochondrial NTP/dNTP ratios. In this chapter, we describe a method for the determination of mtDNA rNMP content using alkaline gel electrophoresis and Southern blotting. This procedure is suited for the analysis of mtDNA in total genomic DNA preparations as well as in purified form. Moreover, it can be performed using equipment found in most biomedical laboratories, allows the simultaneous analysis of 10-20 samples depending on the gel system employed, and can be modified for the analysis of other mtDNA modifications.

sted, utgiver, år, opplag, sider
New York: Humana Press, 2023. Vol. 2615, s. 293-314
Serie
Methods in Molecular Biology, ISSN 1064-3745, E-ISSN 1940-6029 ; 2615
Emneord [en]
Alkaline gels, Alkaline hydrolysis, Denaturing gels, Ribonucleotides, rNMPs, Southern blot
HSV kategori
Identifikatorer
URN: urn:nbn:se:umu:diva-205504DOI: 10.1007/978-1-0716-2922-2_21PubMedID: 36807800Scopus ID: 2-s2.0-85148677775ISBN: 9781071629215 (tryckt)ISBN: 9781071629222 (digital)OAI: oai:DiVA.org:umu-205504DiVA, id: diva2:1743462
Tilgjengelig fra: 2023-03-15 Laget: 2023-03-15 Sist oppdatert: 2023-07-11bibliografisk kontrollert

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