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Monitoring Bacterial Genetic Diversity in a Freshwater Lake Using TTGE and DNA Sequence Analysis

sterile scalpel, eluted by passive diffusion into 1.25x TAE and sequenced.


Results and Discussion
TTGE analysis of freshwater Nitrosospira 16S rDNA PCR products revealed both genotypic continuity and variation between individual samples (Figure 1). Co-migrating DNA bands (position B) were observed during the periods of lakewater overturn and stratification, suggesting the presence of a predominant Nitrosospira genotype during these times. Spring and autumn samples, corresponding to formation and breakdown of a stratified water column respectively, demonstrated the predominance of other strains of Nitrosospira by the observation of differentially migrating PCR products (positions A, C, D, E and F). While myriad other DNA bands present throughout the gel indicated an overall complex genetic profile within this Nitrosospira group, the relative quantity of DNA in individual bands suggested the numerical dominance by a number of strains (positions A to F) whose prevalence appeared to be temporally variable. Six DNA bands (positions A to F) were excised from the gel and sequenced. Phylogenetic analysis of these sequences grouped them as closely-related genotypes, but in a separate cluster were grouped to previously recognized Nitrosospira spp.4

Confidence in dendrogram topology is limited due to the restricted quantity of sequence data available for scrutiny (approx. 200 bp), yet the acute resolving power of the TTGE technique enabled the separation of Nitrosospira 16S rRNA genes differing by as little as one base pair across the fragment examined.

TTGE is a reliable technique and a valuable tool for microbial diversity studies. By loadi
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