Donnerstag, 18. September 2014

Optimization of Extraction of Circulating RNAs from Plasma - Enabling Small RNA Sequencing.


Optimization of Extraction of Circulating RNAs from Plasma - Enabling Small RNA Sequencing.
Spornraft M, Kirchner B, Haase B, Benes V, Pfaffl MW, Riedmaier I
PLoS One. 2014 9(9): e107259 - eCollection 2014

There are several protocols and kits for the extraction of circulating RNAs from plasma with a following quantification of specific genes via RT-qPCR. Due to the marginal amount of cell-free RNA in plasma samples, the total RNA yield is insufficient to perform Next-Generation Sequencing (NGS), the state-of-the-art technology in massive parallel sequencing that enables a comprehensive characterization of the whole transcriptome. Screening the transcriptome for biomarker signatures accelerates progress in biomarker profiling for molecular diagnostics, early disease detection or food safety. Therefore, the aim was to optimize a method that enables the extraction of sufficient amounts of total RNA from bovine plasma to generate good-quality small RNA Sequencing (small RNA-Seq) data. An increased volume of plasma (9 ml) was processed using the Qiagen miRNeasy Serum/Plasma Kit in combination with the QIAvac24 Plus system, a vacuum manifold that enables handling of high volumes during RNA isolation. 35 ng of total RNA were passed on to cDNA library preparation followed by small RNA high-throughput sequencing analysis on the Illumina HiSeq2000 platform. Raw sequencing reads were processed by a data analysis pipeline using different free software solutions. Seq-data was trimmed, quality checked, gradually selected for miRNAs/piRNAs and aligned to small RNA reference annotation indexes. Mapping to human reference indexes resulted in 4.8±2.8% of mature miRNAs and 1.4±0.8% of piRNAs and of 5.0±2.9% of mature miRNAs for bos taurus.

Mittwoch, 17. September 2014

qPCR NEWS -- September 2014 -- Hot papers in the field of microRNA and non-coding RNAs


The September 2014 newsletter informs about the latest news in quantitative real-time PCR (qPCR and RT-qPCR), which are compiled and summarised on the Gene Quantification domain. The focus of this newsletter issue is:
If this newsletter is not displayed correctly please use following LINK => qPCRnews.gene-quantification.info

Donnerstag, 24. Juli 2014

"BDQ Young Scientist Poster Awards" at qPCR & NGS 2015


"BDQ Young Scientist Poster Awards" at qPCR & NGS 2015
BDQ-awards.qPCR-NGS-2015.net

The new open access journal Biomolecular Detection and Quantification (BDQ) is proud to sponsor three BDQ Young Scientist Poster Awards at the qPCR & NGS 2015 Event.

The posters will be evaluated by an international jury on the basis of originality of the approach; quality of the work (e.g. appropriate methodology, interpretation of results, conclusiveness); quality of the presentation (e.g. clarity, response to questions); and self-reliance and independence (not one of co-authors, but the most active, if more than one author).

Eligible for the BDQ Young Scientist Poster Award:
The contest is open to students and early career scientists.

Award Presentation
BDQ Editor-in-Chief Stephen Bustin will award the three poster prizes on Wednesday afternoon directly after the lunch break March 25, 2015.

  1. BDQ Young Scientist Poster Award - First Prize
    The winner is awarded 700 Euros and a free submission to BDQ
  2. BDQ Young Scientist Poster Award – Second Prize
    The winner is awarded 200 Euros and a free submission to BDQ
  3. BDQ Young Scientist Poster Award – Third Prize
    The winner is awarded 100 Euros and a free submission to BDQ
Procedure for Application
Authors will be able to express their interest in participating in the BDQ Young Scientist Poster Award contest when submitting their abstract. Note that authors are only able to participate if their paper has been selected as a poster by the conference organizer.
Please register and submit your abstract using the Internet based ConfTool registration and submission platform => registration.qPCR-NGS-2015.net
 

Montag, 21. Juli 2014

Why are reporting guidelines so essential?

Why are reporting guidelines so essential?

» by LGC

We’ve all probably heard the Chinese proverb that it’s not the destination that is important but the journey. Well the same can be said of scientific research: it’s not only the results that matter but the methodology and processes that lead us to them.
Scientist carries out analysis on genetically modified organism using polymerase chain reaction (PCR)OK, so this may be a little facetious but the concept is not and it’s the reason why guidelines for the reporting of scientific experiments have been emerging over the past decade across scientific disciplines – including the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines.
PCR (polymerase chain reaction) is a technique widely used in molecular biology to identify and quantify DNA. PCR works by targeted amplification of DNA by several orders of magnitude to enable identification and measurement of specific sequences. Quantitative polymerase chain reaction (qPCR), also called real-time polymerase chain reaction, is a laboratory technique based on the PCR, which is used to amplify and simultaneously quantify a targeted DNA molecule.
In order to encourage increased transparency in reported data, the MIQE guidelines were developed and published in 2009. But why is this so important and, five years later, what impact have the guidelines had?