A New Role for Yeast Cells in Health and Nutrition: Antioxidant Power Assessment.

As the use of antioxidant compounds in the domains of health, nutrition and well-being is exponentially rising, there is an urgent need to quantify antioxidant power quickly and easily, ideally within living cells. We developed an Anti Oxidant Power in Yeast (AOPY) assay which allows for the quantitative measurement of the Reactive Oxygen Species (ROS) and free-radical scavenging effects of various molecules in a high-throughput compatible format. Key parameters for Saccharomyces cerevisiae were investigated, and the optimal values were determined for each of them. The cell density in the reaction mixture was fixed at 0.6; the concentration of the fluorescent biosensor (TO) was found to be optimal at 64 µM, and the strongest response was observed for exponentially growing cells. Our optimized procedure allows accurate quantification of the antioxidant effect in yeast of well-known antioxidant molecules: resveratrol, epigallocatechin gallate, quercetin and astaxanthin added in the culture medium. Moreover, using a genetically engineered carotenoid-producing yeast strain, we realized the proof of concept of the usefulness of this new assay to measure the amount of ß-carotene directly inside living cells, without the need for cell lysis and purification.

LUCS: a high-resolution nucleic acid sequencing tool for accurate long-read analysis of individual DNA molecules.

Nucleic acid sequence analyses are fundamental to all aspects of biological research, spanning aging, mitochondrial DNA (mtDNA) and cancer, as well as microbial and viral evolution. Over the past several years, significant improvements in DNA sequencing, including consensus sequence analysis, have proven invaluable for high-throughput studies. However, all current DNA sequencing platforms have limited utility for studies of complex mixtures or of individual long molecules, the latter of which is crucial to understanding evolution and consequences of single nucleotide variants and their combinations. Here we report a new technology termed LUCS (Long-molecule UMI-driven Consensus Sequencing), in which reads from third-generation sequencing are aggregated by unique molecular identifiers (UMIs) specific for each individual DNA molecule. This enables in-silico reconstruction of highly accurate consensus reads of each DNA molecule independent of other molecules in the sample. Additionally, use of two UMIs enables detection of artificial recombinants (chimeras). As proof of concept, we show that application of LUCS to assessment of mitochondrial genomes in complex mixtures from single cells was associated with an error rate of 1X10-4 errors/nucleotide. Thus, LUCS represents a major step forward in DNA sequencing that offers high-throughput capacity and high-accuracy reads in studies of long DNA templates and nucleotide variants in heterogenous samples.