ArticleNucleic acids research2016
Adapting capillary gel electrophoresis as a sensitive, high-throughput method to accelerate characterization of nucleic acid metabolic enzymes.
Article in Nucleic acids research, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 38 papers.
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Who cites it
38 citing papers in PubMed, 67 citations in OpenAlex.
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- DNA rehybridization drives product release from Cas9 ribonucleoprotein to enable multiple-turnover cleavage.Nucleic acids research · 2023Article
- Encoding, Decoding, and Rendering Information in DNA Nanoswitch Libraries.ACS synthetic biology · 2023Article
- Application of microfluidic chip electrophoresis for high-throughput nucleic acid fluorescence fragment analysis assays.NAR genomics and bioinformatics · 2023Article
- RNase H-based analysis of synthetic mRNA 5' cap incorporation.RNA (New York, N.Y.) · 2022Article
- Programmable cleavage of linear double-stranded DNA by combined action of Argonaute CbAgo from Clostridium butyricum and nuclease deficient RecBC helicase from E. coli.Nucleic acids research · 2022Article
- Characterization of Cme and Yme thermostable Cas12a orthologs.Communications biology · 2022Article
- Capillary Electrophoresis-Based Functional Genomics Screening to Discover Novel Archaeal DNA Modifying Enzymes.Applied and environmental microbiology · 2022Article
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- High throughput quantification of short nucleic acid samples by capillary electrophoresis with automated data processing.Analytical biochemistry · 2021Article
- E. coli RNase I exhibits a strong Ca2+-dependent inherent double-stranded RNase activity.Nucleic acids research · 2021Article
- Development of a Cas12a-Based Genome Editing Tool for Moderate Thermophiles.The CRISPR journal · 2021Article
- ProfilingFrontiers in molecular biosciences · 2021Article
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Authors and funding
8 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Detailed biochemical characterization of nucleic acid enzymes is fundamental to understanding nucleic acid metabolism, genome replication and repair. We report the development of a rapid, high-throughput fluorescence capillary gel electrophoresis method as an alternative to traditional polyacrylamide gel electrophoresis to characterize nucleic acid metabolic enzymes. The principles of assay design described here can be applied to nearly any enzyme system that acts on a fluorescently labeled oligonucleotide substrate. Herein, we describe several assays using this core capillary gel electrophoresis methodology to accelerate study of nucleic acid enzymes. First, assays were designed to examine DNA polymerase activities including nucleotide incorporation kinetics, strand displacement synthesis and 3'-5' exonuclease activity. Next, DNA repair activities of DNA ligase, flap endonuclease and RNase H2 were monitored. In addition, a multicolor assay that uses four different fluorescently labeled substrates in a single reaction was implemented to characterize GAN nuclease specificity. Finally, a dual-color fluorescence assay to monitor coupled enzyme reactions during Okazaki fragment maturation is described. These assays serve as a template to guide further technical development for enzyme characterization or nucleoside and non-nucleoside inhibitor screening in a high-throughput manner.
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