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Bacterial Diversity Studies Using the 16S rRNA Gene Provide a Powerful Research-Based Curriculum for Molecular Biology Laboratory

    Authors: SARAH M. BOOMER1,*, DANIEL P. LODGE1, BRYAN E. DUTTON1
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    Affiliations: 1: Western Oregon University, Department of Biology, Monmouth, Oregon 97361
    AUTHOR AND ARTICLE INFORMATION AUTHOR AND ARTICLE INFORMATION
    • *Corresponding author. Mailing address: Western Oregon University, Department of Biology, 345 Monmouth Ave., Monmouth, OR 97361. Phone: (503) 838-8209. Fax: (503) 838-8072. E-mail: boomers@wou.edu.
    • Copyright © 2002, American Society for Microbiology. All Rights Reserved.
    Source: J. Microbiol. Biol. Educ. May 2002 vol. 3 no. 1 18-25. doi:10.1128/154288102X14285807655107
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    Abstract:

    We have developed a ten-week curriculum for molecular biology that uses 16S ribosomal RNA genes to characterize and compare novel bacteria from hot spring communities in Yellowstone National Park. The 16S rRNA approach bypasses selective culture-based methods. Our molecular biology course offered the opportunity for students to learn broadly applicable methods while contributing to a long-term research project. Specifically, students isolated and characterized clones that contained novel 16S rRNA inserts using restriction enzyme, DNA sequencing, and computer-based phylogenetic methods. In both classes, students retrieved novel bacterial 16S rRNA genes, several of which were most similar to Green Nonsulfur bacterial isolates. During class, we evaluated student performance and mastery of skills and concepts using quizzes, formal lab notebooks, and a broad project assignment. For this report, we also assessed student performance alongside data quality and discussed the significance, our goal being to improve both research and teaching methods.

Key Concept Ranking

Restriction Fragment Length Polymorphism
0.4429262
Environmental Microbiology
0.4421141
0.4429262

References & Citations

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2. Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K1997Short protocols in molecular biology3rd edJohn Wiley and Sons, IncNew York, N.Y.
3. Boomer SM, Lodge DP, Dutton BE, Pierson B2002Molecular characterization of novel red green nonsulfur bacteria from five distinct hot spring communities in Yellowstone National ParkAppl Environ Microbiol6834635510.1128/AEM.68.1.346-355.200211772644 http://dx.doi.org/10.1128/AEM.68.1.346-355.2002
4. Boomer SM, Pierson BK, Austinhirst R, Castenholz RW2000Characterization of novel bacterio-chlorophyll-a-containing red filaments from alkaline hot springs in Yellowstone National ParkArch Microbiol17415216110.1007/s00203000018911041345 http://dx.doi.org/10.1007/s002030000189
5. Committee on Development of an Addendum to the National Science Education Standards on Scientific Inquiry2000Inquiry and the National Science Education Standards, a guide for teaching and learningNational Academy PressWashington D.C.
6. Hanada S, Takaichi S, Matsuura K, Nakamura KRoseiflexus castenholzii gen. nov., sp. nov., a thermophilic, filamentous, photosynthetic bacterium which lacks chlorosomesInt. J. Syst. Bacteriol.5218793
7. Hugenholtz P, Goebel BM, Pace NR1998Impact of culture-independent studies on the emerging phylogenetic view of bacterial diversityJ Bacteriol180476547749733676
8. Hurst CJ, Crawford RL, Knudsen GR, McInerney MJ, Stetzenbach LD2002Manual of environmental microbiology2nd edASM PressWashington D.C.
9. Pace NR1997A molecular view of microbial diversity and the biosphereScience27673474010.1126/science.276.5313.7349115194 http://dx.doi.org/10.1126/science.276.5313.734
10. Reysenbach AL, Wickham GS, Pace NR1994Phylogenetic analysis of the hyperthermophilic pink filament community in Octopus Spring, Yellowstone National ParkAppl Environ Microbiol60211321197518219
11. Sanger F, Nicklen S, Coulson AR1977DNA sequencing with chain-terminating inhibitorsProc Natl Acad Sci USA745463546710.1073/pnas.74.12.5463271968 http://dx.doi.org/10.1073/pnas.74.12.5463
12. Winfrey MR, Rott MA, Wortman AT1997Unraveling DNA molecular biology for the laboratoryPrentice-Hall, IncUpper Saddle River, N.J.
13. Woese CR1987Bacterial evolutionMicrobiol Rev512212712439888
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/content/journal/jmbe/10.1128/154288102X14285807655107
2002-05-01
2017-07-21

Abstract:

We have developed a ten-week curriculum for molecular biology that uses 16S ribosomal RNA genes to characterize and compare novel bacteria from hot spring communities in Yellowstone National Park. The 16S rRNA approach bypasses selective culture-based methods. Our molecular biology course offered the opportunity for students to learn broadly applicable methods while contributing to a long-term research project. Specifically, students isolated and characterized clones that contained novel 16S rRNA inserts using restriction enzyme, DNA sequencing, and computer-based phylogenetic methods. In both classes, students retrieved novel bacterial 16S rRNA genes, several of which were most similar to Green Nonsulfur bacterial isolates. During class, we evaluated student performance and mastery of skills and concepts using quizzes, formal lab notebooks, and a broad project assignment. For this report, we also assessed student performance alongside data quality and discussed the significance, our goal being to improve both research and teaching methods.

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Figures

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FIG. 1

Student-generated flow chart depicting large-scale plasmid isolation procedures, Unit 1. This component of the prelab assignment comprised 10% of each Unit Lab grade. Evaluation notes in upper right corner are instructors additions.

Source: J. Microbiol. Biol. Educ. May 2002 vol. 3 no. 1 18-25. doi:10.1128/154288102X14285807655107
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Image of FIG. 2

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FIG. 2

Phylogenetic tree using representative class data. The tree was generated using maximum parsimony methods against a dataset of known bacteria (indicated in italics with accession number in parentheses). The bar indicates 10 nucleotide changes. Branch numbers indicate percent support for that branch. Bacterial lineages are indicated in brackets.

Source: J. Microbiol. Biol. Educ. May 2002 vol. 3 no. 1 18-25. doi:10.1128/154288102X14285807655107
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Image of FIG. 3

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FIG. 3

Representative PCR data. Panel A was generated using general bacterial 16S primers. Panel B was generated with GNS-specific primers. Lane 1 in both A and B is marker standards (Lambda/HindIII), and lanes 2 through 12 were representative products using different PCR buffers. The arrows at the right indicate the target fragment. PCR product was separated using 1% agarose with standard TAE running buffer.

Source: J. Microbiol. Biol. Educ. May 2002 vol. 3 no. 1 18-25. doi:10.1128/154288102X14285807655107
Download as Powerpoint

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