Effects of Lyse-It on endonuclease fragmentation, function and activity


Autoři: Tonya M. Santaus aff001;  Fan Zhang aff001;  Shan Li aff003;  O. Colin Stine aff003;  Chris D. Geddes aff001
Působiště autorů: Chemistry and Biochemistry Department, University of Maryland, Baltimore County, Baltimore, Maryland, United States of America aff001;  Institute of Fluorescence, University of Maryland, Baltimore County, Baltimore, Maryland, United States of America aff002;  Epidemiology and Public Health Department, University of Maryland School of Medicine, Baltimore, Maryland, United States of America aff003
Vyšlo v časopise: PLoS ONE 14(9)
Kategorie: Research Article
doi: 10.1371/journal.pone.0223008

Souhrn

Nucleases are enzymes that can degrade genomic DNA and RNA that decrease the accuracy of quantitative measures of those nucleic acids. Here, we study conventional heating, standard microwave irradiation, and Lyse-It, a microwave-based lysing technology, for the potential to fragment and inactivate DNA and RNA endonucleases. Lyse-It employs the use of highly focused microwave irradiation to the sample ultimately fragmenting and inactivating RNase A, RNase B, and DNase I. Nuclease size and fragmentation were determined visually and quantitatively by SDS polyacrylamide gel electrophoresis and the mini-gel Agilent 2100 Bioanalyzer system, with a weighted size calculated to depict the wide range of nuclease fragmentation. Enzyme activity assays were conducted, and the rates were calculated to determine the effect of various lysing conditions on each of the nucleases. The results shown in this paper clearly demonstrate that Lyse-It is a rapid and highly efficient way to degrade and inactivate nucleases so that nucleic acids can be retained for down-stream detection.

Klíčová slova:

Oxygen – Reactive oxygen species – Vibrio cholerae – Nucleases – Ribonucleases – Microwave radiation – Deoxyribonucleases – SDS polyacrylamide gel electrophoresis


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PLOS One


2019 Číslo 9

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