Introduction of green fluorescent protein gene into phenol-degrading Alcaligenes faecalis cells and their monitoring in phenol-contaminated soil

Abstract

Alcaligenesfaecalis (CCT 7145) was isolated from an Amazonian soil sample after an enrichment process to select for phenol-degrading microorganisms. The isolate was labeled with the green fluorescent protein (gfp) gene. The gfp-transformed cells were easily detected using a hand-held UV transilluminator and their taxonomy was confirmed by 16S rRNA sequencing. Polymerase chain reaction (PCR) and Southern blot analyses confirmed that the gfp gene was integrated into the chromosome. The addition of the gfp marker did not affect phenol degradation ability compared with the wild-type. Both, wild-type and gfp-marked A. faecalis cells encapsulated in alginate, tolerated 1,700 microg ml(-1) phenol in liquid medium compared with 1,100 microg ml(-1) phenol for free cells. 14C-Phenol mineralization in soil microcosms was also enhanced by inoculation with encapsulated cells. Survival of gfp-marked cells in phenol-contaminated soil over 22 days was determined from plate counts using an epifluorescence microscope.

A safety type genetically engineered bacterium with red fluorescence which can be used to degrade organophosphorus pesticides

“…To some extent, this result confirms previous studies which showed that fluorescence can be used to monitor GEMs (Leff and Leff 1996;Bastos et al 2001).…”

New Approaches for Bioaugmentation as a Remediation Technology

“…Alginate is the most commonly investigated carrier for bioremediation applications, and has been used with numerous contaminants including chromium, cresol, nitrate, pentachlorophenol, phenanthrene, phenol, phosphate, and 2,4,6-trichlorophenol. 10,28,73,75,87,141,166,206,250 Alginate may also have potential for delivery of naked DNA directly into the environment for the purpose of gene bioaugmentation, 1 which is discussed in a later section.…”

“…The capsule matrix can buffer the introduced microorganisms against pollutant toxicity in the environment. 10,149 Additionally, substrates or C compounds can be added to the capsule to confer an advantage to the embedded inoculant; 236 however, Duquenne et al…”

“…10,43 The GFP-containing cells can be detected by epifluorescence microscopy or by visualization of cultured bacteria on plates with long-wave ultraviolet (UV) or black light. 10,237 The gfp-tagged cells can also be used to detect gene expression. 34,96 The gfp gene has found particular use in dual-labeling applications with other reporter genes.…”

Slow-Release Inoculation Allows Sustained Biodegradation of γ-Hexachlorocyclohexane

“…The capsule matrix can buffer against environmental stress and pollutant toxicity (5,23) and protects the inoculum from predation and indigenous microorganisms (29) while allowing the diffusion of gases and liquids (37). Carrier materials to encapsulate cells for biodegradation include, among others, alginate, gelatin, and -carrageenan gel (36).…”