High-Throughput Screening Platform for Longevity Genes and Anti-Aging Drugs
AssigneeThe Regents of the University of California
InventorsChanghui Deng, Hao Li
Life of this patent
Estimating the term…
Abstract
Compositions, devices, and systems for use in a high-throughput screening platform for identifying anti-aging compounds and/or mutations that extend replicative life span (RLS). Specifically, herein disclosed is a yeast cell daughter-arresting-program (DAP), as well as compositions used in devices and systems that allow measurement of replicative lifespan and identification of agents or mutations that modulate the lifespan.
Claims78
A nucleic acid construct for integration into a specific locus of a yeast cell genome, comprising:
- (a) an integration sequence at each end of the nucleic acid construct configured to effect integration into a yeast genomic locus between a sequence upstream of the start codon of an endogenous gene encoding an essential plasma membrane protein and the start codon of the gene;
- and
- (b) two cassettes oriented in opposite transcriptional directions, comprising:
- (i) a first cassette comprising a mother-specific promoter configured to control transcription of an exogenous copy of the gene encoding the essential plasma membrane protein;
- and
- (ii) a second cassette comprising a conditional promoter configured to control transcription of the endogenous gene upon integration into the yeast genomic locus.
A daughter-arresting program (DAP) yeast strain, comprising:
- an exogenous nucleic acid sequence integrated into the genome between a sequence upstream of the start codon of an endogenous gene encoding an essential plasma membrane protein and the start codon of the gene, wherein the integrated nucleic acid sequence comprises:
- (a) a mother-specific promoter driving transcription of an exogenous copy of the gene encoding the essential plasma membrane protein;
- and
- (b) a conditional promoter driving transcription of the endogenous gene encoding the essential plasma membrane protein, wherein the mother-specific promoter and the conditional promoter are oriented in opposite transcriptional directions.
A microfluidic device comprising a plurality of functional modules for measurement of yeast replicative lifespan (RLS), wherein each module comprises:
- (a) an inlet for receiving fluid flow into the module,
- (b) a cell-trapping and observational area, in fluid communication with the inlet, comprising an array of trapping units configured to trap budding mother cells and arrested daughter cells produced therefrom, and
- (c) an outlet, in fluid communication with the cell-trapping and observational area, for flow out of the module.
A yeast cell culture device comprising a multiwell plate integrated with a microfluidic device positioned beneath the multiwell plate, the microfluidic device comprising a plurality of functional modules for measurement of RLS, wherein each module corresponds to a plurality of wells of the multiwell plate, and wherein each module comprises:
- (a) an inlet configured to provide fluid flow into the module from a first well of the multiwell plate,
- (b) a cell-trapping and observational area in fluid communication with the inlet and comprising an array of trapping units for trapping budding mother cells and arrested daughter cells produced therefrom, and
- (c) an outlet in fluid communication with the cell-trapping and observational area, configured to provide fluid flow out of the module to a second well of the multiwell plate.
Dependencies are inferred from the claim text.