Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples
Extraction procedures for RNA and DNA in crude extracts of natural microplankton samples are developed. The methodology is compatible with the fluorometric assay of RNA and DNA developed by Berdalet et al. (2005). Mechanical cell disruption using a manual tissue grinder is combined with chemical ext...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
| Published: |
Consejo Superior de Investigaciones Científicas
2005-03-01
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| Series: | Scientia Marina |
| Subjects: | |
| Online Access: | http://scientiamarina.revistas.csic.es/index.php/scientiamarina/article/view/231 |
| _version_ | 1818863701336260608 |
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| author | Elisa Berdalet Cristina Roldán M. Pilar Olivar |
| author_facet | Elisa Berdalet Cristina Roldán M. Pilar Olivar |
| author_sort | Elisa Berdalet |
| collection | DOAJ |
| description | Extraction procedures for RNA and DNA in crude extracts of natural microplankton samples are developed. The methodology is compatible with the fluorometric assay of RNA and DNA developed by Berdalet et al. (2005). Mechanical cell disruption using a manual tissue grinder is combined with chemical extraction using 0.5% sarcosine and 0.5 mM EDTA in 5 mM Tris, pH 8.0, to release the nucleic acid. The new extraction and assay procedure is used to estimate the RNA/DNA ratios of fish larvae and natural microplankton communities maintained in the laboratory under different nutritional conditions. In the two experiments, the RNA/DNA ratios were related to the nutrient availability of the organisms. The level of sensitivity of the method is experimentally set at ca. 40 ng of RNA and 10 ng DNA in the 1 ml assay, which corresponds to a minimum biomass requirement of ca. 400 ng ml-1 protein or 3 µg ml-1 dry weight. The precision, estimated at different steps of the procedure, had an overall coefficient of variation of ?10%. The new approach provides RNA/DNA ratios comparable with those obtained with ethidium bromide. |
| first_indexed | 2024-12-19T10:19:56Z |
| format | Article |
| id | doaj.art-77dc8227aa30465281be3c72a65e09a5 |
| institution | Directory Open Access Journal |
| issn | 0214-8358 1886-8134 |
| language | English |
| last_indexed | 2024-12-19T10:19:56Z |
| publishDate | 2005-03-01 |
| publisher | Consejo Superior de Investigaciones Científicas |
| record_format | Article |
| series | Scientia Marina |
| spelling | doaj.art-77dc8227aa30465281be3c72a65e09a52022-12-21T20:26:04ZengConsejo Superior de Investigaciones CientíficasScientia Marina0214-83581886-81342005-03-01691173010.3989/scimar.2005.69n117228Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samplesElisa Berdalet0Cristina Roldán1M. Pilar Olivar2Institut de Ciències del Mar (CSIC)Institut de Ciències del Mar (CSIC)Institut de Ciències del Mar (CSIC)Extraction procedures for RNA and DNA in crude extracts of natural microplankton samples are developed. The methodology is compatible with the fluorometric assay of RNA and DNA developed by Berdalet et al. (2005). Mechanical cell disruption using a manual tissue grinder is combined with chemical extraction using 0.5% sarcosine and 0.5 mM EDTA in 5 mM Tris, pH 8.0, to release the nucleic acid. The new extraction and assay procedure is used to estimate the RNA/DNA ratios of fish larvae and natural microplankton communities maintained in the laboratory under different nutritional conditions. In the two experiments, the RNA/DNA ratios were related to the nutrient availability of the organisms. The level of sensitivity of the method is experimentally set at ca. 40 ng of RNA and 10 ng DNA in the 1 ml assay, which corresponds to a minimum biomass requirement of ca. 400 ng ml-1 protein or 3 µg ml-1 dry weight. The precision, estimated at different steps of the procedure, had an overall coefficient of variation of ?10%. The new approach provides RNA/DNA ratios comparable with those obtained with ethidium bromide.http://scientiamarina.revistas.csic.es/index.php/scientiamarina/article/view/231sybr green iidnasernasesarcosinerna/dna ratiosplankton |
| spellingShingle | Elisa Berdalet Cristina Roldán M. Pilar Olivar Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples Scientia Marina sybr green ii dnase rnase sarcosine rna/dna ratios plankton |
| title | Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples |
| title_full | Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples |
| title_fullStr | Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples |
| title_full_unstemmed | Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples |
| title_short | Quantifying RNA and DNA in planktonic organisms with SYBR Green II and nucleases. Part B. Quantification in natural samples |
| title_sort | quantifying rna and dna in planktonic organisms with sybr green ii and nucleases part b quantification in natural samples |
| topic | sybr green ii dnase rnase sarcosine rna/dna ratios plankton |
| url | http://scientiamarina.revistas.csic.es/index.php/scientiamarina/article/view/231 |
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