Isolation of Morphological variants in Variovorax paradoxus EPS for genome sequencing
Poster Number
8C
Research or Creativity Area
Natural Sciences
Abstract
Variovorax paradoxus is a species of bacteria usually found in soil, and known to positively influence plant growth as a component of the rhizosphere. Plant growth promotion in crops and bioremediation by V. paradoxus are important potential applications of research on this bacterium. Th EPS strain of this bacterium was cultivated for its copious production of expolysaccharides. The purpose of the current research is to evaluate a whole genome sequencing approach to identifying genes that contribute to colony morphology. The goal is to isolate multiple morphological variants that “breed true” on plates, and use whole genome resequencing to identify genes that contribute to this complex trait. We will compare them to their reference genomes to identify mutations in the colony morphology variants, and by looking at independent isolates with similar phenotypes, we will aim to find genes that are critical to the colony development process.
Growth medium was made using the Freshwater medium previously described, with 2 mM sodium succinate as the sole carbon source. The medium as buffered with 5 mM MOPS to maintain a pH around 7. Overnight cultures of V. paradoxus EPS were grown in 5 g/L yeast extract broth. The cultures were washed in water and then dilution plated onto the FW-succinate plates. Agarose was used as the solidifying agent in the medium to improve reproducibility. Cultures were grown at 30 degrees Celcius and colonies were chosen based on morphology. Fourteen isolates were restreaked on the same medium to verify the stability of the trait. Stock cultures of each isolate are generated, and DNA is extracted for sequencing using the Promega Wizard genomic kit. DNA sequencing using the Oxford Nanopore MinION single molecule instrument is performed using the Rapid barcoding kit that allows 24 samples to be sequenced on a single flow cell. The isolate genomes will be assembled using Trycycler and compared to the reference genome.
Location
Don and Karen DeRosa University Center (DUC) Poster Hall
Start Date
27-4-2024 10:30 AM
End Date
27-4-2024 12:30 PM
Isolation of Morphological variants in Variovorax paradoxus EPS for genome sequencing
Don and Karen DeRosa University Center (DUC) Poster Hall
Variovorax paradoxus is a species of bacteria usually found in soil, and known to positively influence plant growth as a component of the rhizosphere. Plant growth promotion in crops and bioremediation by V. paradoxus are important potential applications of research on this bacterium. Th EPS strain of this bacterium was cultivated for its copious production of expolysaccharides. The purpose of the current research is to evaluate a whole genome sequencing approach to identifying genes that contribute to colony morphology. The goal is to isolate multiple morphological variants that “breed true” on plates, and use whole genome resequencing to identify genes that contribute to this complex trait. We will compare them to their reference genomes to identify mutations in the colony morphology variants, and by looking at independent isolates with similar phenotypes, we will aim to find genes that are critical to the colony development process.
Growth medium was made using the Freshwater medium previously described, with 2 mM sodium succinate as the sole carbon source. The medium as buffered with 5 mM MOPS to maintain a pH around 7. Overnight cultures of V. paradoxus EPS were grown in 5 g/L yeast extract broth. The cultures were washed in water and then dilution plated onto the FW-succinate plates. Agarose was used as the solidifying agent in the medium to improve reproducibility. Cultures were grown at 30 degrees Celcius and colonies were chosen based on morphology. Fourteen isolates were restreaked on the same medium to verify the stability of the trait. Stock cultures of each isolate are generated, and DNA is extracted for sequencing using the Promega Wizard genomic kit. DNA sequencing using the Oxford Nanopore MinION single molecule instrument is performed using the Rapid barcoding kit that allows 24 samples to be sequenced on a single flow cell. The isolate genomes will be assembled using Trycycler and compared to the reference genome.
