Following their initial work, scientists began analyzing the genomes of various organisms known for their ability to degrade industrial pollutants. The process of breaking down complex molecules often requires multiple enzymes, which are typically encoded by genes that cluster together. This genetic arrangement allows for the synthesis of a large RNA molecule encompassing all necessary proteins, thereby simplifying the regulation of their production. The clusters identified in this research varied in size, containing between three to eleven genes.
After pinpointing nine such gene clusters, the research team ordered the corresponding DNA sequences and assembled them into a singular DNA molecule using yeast as a host. During this ordering process, they also optimized the genes to ensure high activity and protein production specifically in Vibrio natriegens, rather than the original host organisms for these genes.
The individual gene clusters were then introduced into Vibrio natriegens, resulting in different bacterial strains engineered to metabolize compounds including benzene, toluene, phenol, naphthalene, biphenyl, DBF29, and dibenzothiophene (DBT). Several of the nine identified clusters were designed to target the same contaminants. The modified bacterial strains were subsequently tested in solutions containing the respective chemicals, with five out of the nine demonstrating effective degradation capabilities. The successful strains could break down biphenyl, phenol, naphthalene, DBF, and toluene.
Progress with Limitations
The researchers then devised a method to incrementally insert a new gene cluster at the end of an already integrated cluster. This innovative approach allowed them to accumulate a ‘cluster of clusters,’ ultimately including all five strains demonstrated to be effective in prior experiments. After two days of operation, this composite strain could eliminate approximately 25% of phenol, 33% of biphenyl, 30% of DBF, all of the naphthalene, and nearly all of the toluene from the solutions.
