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Mapping Ribonucleotides Incorporated into DNA by Hydrolytic End-Sequencing.
Clinton D Orebaugh1, Scott A Lujan1, Adam B Burkholder2
1Genome Integrity and Structural Biology Laboratory, National Institute for Environmental Health Sciences, National Institute of Health (NIH), 111 TW Alexander Drive, Research Triangle Park, NC, 27709, USA.
Methods in Molecular Biology (Clifton, N.J.)
|October 19, 2017
Summary
This study introduces hydrolytic end sequencing (HydEn-seq) to map ribonucleotides in yeast DNA. The method reveals how these embedded ribonucleotides affect DNA stability and genomic features.
Area of Science:
- Molecular Biology
- Genomics
- Biochemistry
Background:
- Ribonucleotides can be mistakenly incorporated into DNA during replication.
- Incorporated ribonucleotides compromise DNA integrity, leading to strand breaks under alkaline conditions.
- Efficient repair mechanisms are crucial for removing these ribonucleotides.
Purpose of the Study:
- To develop and apply a novel sequencing method for mapping ribonucleotides in the genome.
- To investigate the genomic distribution of ribonucleotides in yeast strains with varying repair capacities.
- To understand the impact of ribonucleotides on DNA stability and genomic features.
Main Methods:
- Development of hydrolytic end sequencing (HydEn-seq), a next-generation sequencing technique.
- Application of HydEn-seq to map ribonucleotides in Saccharomyces cerevisiae strains.
- Analysis of genomic features in wild-type and replicase variant yeast strains.
Main Results:
- HydEn-seq successfully mapped embedded ribonucleotides within the yeast genome.
- The study identified genomic locations sensitive to ribonucleotide incorporation.
- Differences in ribonucleotide mapping were observed between wild-type and variant yeast strains.
Conclusions:
- HydEn-seq is a powerful tool for analyzing ribonucleotide incorporation and its genomic consequences.
- Understanding ribonucleotide dynamics is essential for genome stability research.
- This method provides insights into DNA repair pathways and potential therapeutic targets.