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A laboratory unit investigating 2,4-dichlorophenoxyacetic acid degradation in soils

A laboratory unit investigating 2,4-dichlorophenoxyacetic acid degradation in soils Teresa Johnson, College of Wooster Amy Treonis, University of Richmond Diuto Esiobu, Florida Atlantic University. Sample 2,4-D degradation pathway. Who degrades 2,4-D in soils?. Pseudomonas spp.

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A laboratory unit investigating 2,4-dichlorophenoxyacetic acid degradation in soils

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  1. A laboratory unit investigating 2,4-dichlorophenoxyacetic acid degradation in soils Teresa Johnson, College of Wooster Amy Treonis, University of Richmond Diuto Esiobu, Florida Atlantic University

  2. Sample 2,4-D degradation pathway

  3. Who degrades 2,4-D in soils? Pseudomonas spp. Arthrobacter spp.

  4. Audience: Biology Majors in a Microbiology course (intermediate to upper level) Objectives: • Students who complete these activities will: • Explore environmental problems using bioinformatics tools. • Appreciate metabolic flexibility of microbes through study of their ability to use novel chemicals (xenobiotics) as energy sources. • Integrate concepts in functional genomics, phylogenetics, and ecology.

  5. Step 1: Finding 2,4-D degraders • Challenge students to develop an appropriate enrichment culture technique & isolate a 2,4-D degrader in pure culture.

  6. Step 2: Identification of isolate • 16S rDNA sequencing from pure culture • BLAST search to identify microbe

  7. Step 3: Functional genomics • BLAST search for tfdA amino acid sequences (Pseudomonas) • Use aa sequence from BLAST to perform a JGI search • for sequenced bacteria that contain the gene to determine whether our isolate has been identified as having tfdA genes. • Design primers to attempt to amplify tfdA genes from isolate • for sequencing. • If tfdA genes aren’t present, propose alternative mechanism • for degradation. • If tfdA genes are present, compare aa or nucleotide sequence similarity between known degraders and our isolate.

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