Observation Hypothesis Experimental Design (including Methods) Results Inference Camp Wildness 2004 Ward Lab Research Project.

Observation

Hypothesis

Experimental Design (including Methods)

Results

Inference

Camp Wildness 2004 Ward Lab Research Project

The concept of housekeeping genes

ATPase

DNA-dependantDNA polymerase

DNA-dependantRNA polymerase

ribosomecomponents

Ribosome components

Small subunit (16S) rRNA

Highly variablesegment

Highly conservedsegment

PCR amplification of SSU rRNA genes

Focus on a particular gene in the genome

Extract DNA

PCR

Mixed 16S rRNAgenes

Molecular analysisof microbial communities

PCR amplificationOf 16S rRNA gene

Agarose gel analysis of PCR product

Separate by DGGE

Increasing D

enaturant

Mixed Population DNAs

+16S rRNA Gene G+C “Clamp”

PCR Primers G+C-Tailed ProductSeparate onDenaturingGradient

Gel

PCR Amplification

Denaturing Gradient Gel Electrophoresis

Separation Based onDifferences in

Nucleotide Sequence (G+C content) and Melting

Characteristics

B CMix A

Purified Bands for Sequence Analysis

DGGE analysis of 16S rRNA sequences

DGGE study of temperature distribution of Octopus Springcyanobacterial mat 16S rRNA variants

ecotype

Extract DNA

PCR

Native microbialpopulations

Mixed 16S rRNA

Separate by DGGE

Sequencing

Analyzing distributionof molecular diversity

Denaturing GradientGel Electrophoresis

Phylogenetic identification

Gloeobacter violaceus

Pseudoanabaena galeata

OS Type P Isolate

Oscillatoria limnetica

Oscillatoria sp. 7515

Microcoleus sp.

OS C1 Isolate

Synechococcus sp. 6301

Oscillatoria sp. 6304

Oscillatoria amphigranulata

OS Type I

Spirulina sp.

Anabaena cylindrica

Pleurocapsa sp.

Synechocystis sp.

Phormidium ectocarpi

OS C9 Isolate

0.01

OS Type J

OS Type B Isolate

OS Type B' Isolate

OS Type A''

OS Type A'

OS Type A

OS Type A'''

Yellowstone cyanobacterialmat cyanobacterial 16S rRNAdiversity

Readily cultured

predominant in situ

A natural view of microbiology

Observation

Hypothesis

Experimental Design (including Methods)

Results

Inference

Camp Wildness 2004 Ward Lab Research Project