Connecting Microbial Genotype with Phenotype in the Omics Era.

Area of Science:

• Microbiology
• Biotechnology
• Synthetic Biology

Background:

• Rational design is common in metabolic engineering, but conventional methods like adaptive laboratory evolution (ALE) and mutagenesis remain effective for enhancing microbial capabilities.
• These traditional approaches are crucial for developing biocatalysts with improved substrate utilization, growth robustness, and higher product titers, yields, and productivity.

Purpose of the Study:

• To detail methods for generating mutant microbial strains with desirable phenotypes using ALE and Atmosphere and Room Temperature Plasma (ARTP) mutagenesis.
• To outline a strategy for characterizing these mutants by integrating next-generation sequencing (NGS)-based genome resequencing and RNA-Seq transcriptomics.

Main Methods:

• Adaptive Laboratory Evolution (ALE) for strain improvement.
• Atmosphere and Room Temperature Plasma (ARTP) mutagenesis for generating genetic diversity.
• Next-generation sequencing (NGS) for whole-genome resequencing.
• RNA-Seq transcriptomics for analyzing gene expression profiles.

Main Results:

• Successful generation of mutant strains exhibiting enhanced phenotypes through ALE and ARTP.
• Establishment of a combined NGS and RNA-Seq methodology for comprehensive mutant characterization.
• Correlation of observed phenotypic improvements with specific genotypic alterations.

Conclusions:

• ALE and ARTP are powerful tools for microbial strain improvement, complementing rational design.
• Integrating genomic and transcriptomic analyses provides deep insights into genotype-phenotype relationships in engineered microbes.
• This combined approach accelerates the development of superior microbial biocatalysts for industrial applications.