Gene mapping Using Microarrays for PCB Biodegradation

Howard University, Washington DC

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Abstract

Human Health hazards of PCB's (polychlorobiphenyls) are well documented. PCBs enter the human body through lungs and gastro-intestinal tract via food. These can cause reproductive, neurobiological and immunological defects. An understanding of biodegradation pathways of PCBs is not complete until genes and enzymes involved are identified. PCB induced altered gene expressions related to various human health problems are known, but not so for gene expressions involved in PCB biodegradation. Our past experience has raised involved questions such as: (1) Why certain toxic metabolites are not seen when unknown mixed cultures from contaminated soil/sediments are used? (2). What are the pools of genes involved in plant rhizosphere? (3) Do different PCB contaminated sites provide more pool of genes or presence of interacting genes? Based on publications, our hypothesis is that PCB can induced specific gene expressions. Specific Aims of this project are I: Assessments of soils/sediments collected by use from diverse sites of PCB degradation under anaerobic and aerobic conditions; Aim II: Gene array Analysis: Single Vs. mixed cultures, and Aim III: Over expression of candidate genes. Availability of commercially prepared genomic microarrays of E. coli, yeast, and recently sequenced N2-fixing Sinorhizobium sp. Will enable us to hybridize fluorescent labeled total mRNA's isolated from PCB treated cells grown under different conditions. Candidate genes showing over-expressing in PCB treated cells will be subcloned in suitable expression vectors and transformed into E. coli cells. These studies will be repeated using combination of known and unknown single and mixed cultures with or without symbiotic luguminous plants. These retooled studies will enhance significant measurable knowledge regarding (1) effective degradation of PCBs and (2) powerful gene array analysis will enhance new knowledge for PCB induced gene expression which should enhance our understanding of biochemical pathway(s) in PCB biodegradation.