Production and turnover of viruses and dissolved DNA pools at station ALOHA: potential effects on bacteria and roles in the phosphorus cycle
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2003-05
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University of Hawaii at Manoa
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A novel method was developed for the quantification ofdissolved DNA (D-DNA) in seawater. This method includes addition of tetrasodium EDTA to 0.22 um-filtered seawater, concentration of >10kDa material in the filtrate with a Centricon centrifugal ultrafiltration unit, and quantification of the concentrated D-DNA with the fluorescent DNA stain SYBR Green I. This method requires only 13.5 ml of seawater per sample even in ultraoligotrophic environments and samples can be analyzed in less than 3 hours. The recovery of D-DNA with this method is 75-85% and can be determined for each sample by measuring recovery of 35S-labeled DNA added at trace amounts. This method can be used to quantify D-DNA concentrations as low as 0.01 ng ml-1 with high precision (standard deviation <5% of the mean). Deoxyribonuclease (DNase) treatment of samples and virus enumeration can be used in conjunction with this method to determine the three major pools of D-DNA: free or soluble DNA (the fraction hydrolyzable by DNase), DNA within viruses, and uncharacterized bound DNA.
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ix, 73 leaves
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Theses for the degree of Master of Science (University of Hawaii at Manoa). Oceanography; no. 3765
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