Diversity of Expressed Biodegradation Genes in a Contaminated Field SiteEPA Grant Number: U915386
Title: Diversity of Expressed Biodegradation Genes in a Contaminated Field Site
Investigators: Bakermans, Corien M.
Institution: Cornell University
EPA Project Officer: Just, Theodore J.
Project Period: August 1, 1998 through July 1, 2001
Project Amount: $90,879
RFA: STAR Graduate Fellowships (1998) RFA Text | Recipients Lists
Research Category: Fellowship - Microbiology , Biology/Life Sciences , Academic Fellowships
The objectives of this research project are to: (1) explore the diversity of naphthalene dioxygenase (nahAc) genes at a coal tar waste contaminated field site; and (2) determine the contributions of different alleles to naphthalene degradation at the field site.
Both culture-based and molecular approaches will be used to assess the diversity of nahAc genes at the field site to determine how method-specific biases can affect interpretation of diversity. Naphthalene degrading organisms will be isolated from soil and groundwater samples. These isolates will be examined by hybridization, polymerase chain reaction, and cloning to determine the relatedness of their naphthalene degradation genes to the model pathway. To analyze genes from organisms that may not be cultivable by the techniques used, nucleic acids will be extracted directly from site soil and water and analyzed as above. The extraction of DNA will permit the examination of the gene pool present at the site, while extraction of mRNA will enable the analysis of genes that are being actively transcribed (expressed). A phylogenetic analysis of the nahAc genes will allow a comparative analysis of nucleic acids obtained via the direct extraction and the cultivation procedures. If sequences obtained using these two distinct procedures are not closely related, it may be concluded that pure culture techniques were not adequate for assessing the diversity of nahAc genes in the field site. The contribution made by nahAc alleles to degradation of naphthalene at the field site will be assessed by identifying those organisms capable of degrading naphthalene (using substrate-specific direct viable counts) and that express nahAc alleles (using fluorescent in situ hybridization). Together, the two techniques can identify which nahAc alleles are being used for naphthalene degradation at the site.