dc.contributor.author | Tatti, Enrico | |
dc.contributor.author | McKew, Boyd A. | |
dc.contributor.author | Whitby, Corrine | |
dc.contributor.author | Smith, Cindy J. | |
dc.date.accessioned | 2018-09-20T16:26:22Z | |
dc.date.available | 2018-09-20T16:26:22Z | |
dc.date.issued | 2016-06-11 | |
dc.identifier.citation | Tatti, Enrico; McKew, Boyd A. Whitby, Corrine; Smith, Cindy J. (2016). Simultaneous dna-rna extraction from coastal sediments and quantification of 16s rrna genes and transcripts by real-time pcr. Journal of Visualized Experiments (112), | |
dc.identifier.issn | 1940-087X | |
dc.identifier.uri | http://hdl.handle.net/10379/14123 | |
dc.description.abstract | Real Time Polymerase Chain Reaction also known as quantitative PCR (q-PCR) is a widely used tool in microbial ecology to quantify gene abundances of taxonomic and functional groups in environmental samples. Used in combination with a reverse transcriptase reaction (RT-q-PCR), it can also be employed to quantify gene transcripts. q-PCR makes use of highly sensitive fluorescent detection chemistries that allow quantification of PCR amplicons during the exponential phase of the reaction. Therefore, the biases associated with 'end-point' PCR detected in the plateau phase of the PCR reaction are avoided. A protocol to quantify bacterial 16S rRNA genes and transcripts from coastal sediments via real-time PCR is provided. First, a method for the co-extraction of DNA and RNA from coastal sediments, including the additional steps required for the preparation of DNA-free RNA, is outlined. Second, a step-by-step guide for the quantification of 16S rRNA genes and transcripts from the extracted nucleic acids via q-PCR and RT-q-PCR is outlined. This includes details for the construction of DNA and RNA standard curves. Key considerations for the use of RT-q-PCR assays in microbial ecology are included. | |
dc.publisher | MyJove Corporation | |
dc.relation.ispartof | Journal of Visualized Experiments | |
dc.rights | Attribution-NonCommercial-NoDerivs 3.0 Ireland | |
dc.rights.uri | https://creativecommons.org/licenses/by-nc-nd/3.0/ie/ | |
dc.subject | environmental sciences | |
dc.subject | issue 112 | |
dc.subject | DNA-rna co-extraction | |
dc.subject | rna preparation | |
dc.subject | sediment | |
dc.subject | 16s rrna gene | |
dc.subject | 16s rrna transcripts | |
dc.subject | q-pcr | |
dc.subject | rt-q-pcr | |
dc.subject | real-time pcr | |
dc.subject | soil | |
dc.subject | diversity | |
dc.subject | community | |
dc.subject | denitrification | |
dc.subject | microorganisms | |
dc.subject | amplification | |
dc.subject | polymerase | |
dc.subject | recovery | |
dc.subject | bacteria | |
dc.subject | samples | |
dc.title | Simultaneous dna-rna extraction from coastal sediments and quantification of 16s rrna genes and transcripts by real-time pcr | |
dc.type | Article | |
dc.identifier.doi | 10.3791/54067 | |
dc.local.publishedsource | https://www.jove.com/pdf/54067/simultaneous-dna-rna-extraction-from-coastal-sediments-quantification | |
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