Coupling of Realistic Rate Estimates with Genomics for Assessing Contaminant Attenuation and Long-Term Plume Containment [report]

Roland Hirsch
2003 unpublished
Objective. Mr. Erwin's primary objective is to create a non-culture based metagenomic library of the attached and planktonic microbial communities of the Snake River Plain Aquifer (SRPA). This library will then be probed for the presence of genes in the microbial communities that may be used in the cometabolism of chlorinated aliphatics such as trichloroethylene (TCE). Methods. Our approach to this objective includes four phases: sample collection, DNA processing, clone production, and library
more » ... creening. Samples are collected for both attached and planktonic communities existing within the SRPA. The attached organisms are collected by suspending sterile substrate columns (basalt coupons) into the aquifer for a period of 3-6 months. Planktonic organisms are collected via hollow fiber filtration. Metagenomic DNA is obtained from the resulting biomass of each collection through a lysozyme/hot SDS treatment. Gel electrophoresis is then used to isolate 25-40 kb fragments of the randomly sheared DNA. These fragments are electroeluted, end repaired for ligation into Fosmid vectors, and used in the production of our metagenomic library. The libraries are then screened by macro-array analysis for genomic markers of such enzymes as methane monooxygenase and other alkane monooxygenases present in the indigenous microorganisms. Figure 1 shows one of our field sampling campaigns in progress (water filtration) at the INEEL. Figure 2 shows a closeup of the coupon apparatus used to contain basalt chips within the aquifer during growth of biofilms. Figure 1. Sampling of groundwater microbial populations by filtration at the INEEL.
doi:10.2172/836443 fatcat:hwy4ehzt7fae3h37pd5txa5u7i