qPCR NEWS - June 2011 - focus on RNA integrity

Editor www.Gene-Quantification.info via methods%40net.bio.net (by editor from gene-quantification.info)
Mon Jun 27 10:05:54 EST 2011

qPCR NEWS - June 2011 - focus on RNA integrity

Dear researcher,
dear Gene Quantification page reader,

Our newsletter informs about the latest news in quantitative real-time
PCR (qPCR and qRT-PCR), which are compiled and summarised on the Gene
Quantification homepage. The focus of this newsletter issue is:

- Update  -  RNA integrity  -  http://RNA-integrity.gene-quantification.info
- qPCR 2011 eConference  -  http://eConference.qPCR2011.net
- US qPCR Symposium in Santa Clara, CA - 1st-4th November 2011  -
- MIQE qPCR APP for iPhone, iPad and iPod - iOS Universal  -



It is generally known that total RNA quality has a distinct influence
on the validity and reliability of quantitative PCR results. In
addition, the recently published MIQE guidelines focus on the pre-PCR
steps and state the importance of RNA quality assessment. Various
studies showed the impairing effect of ongoing RNA degradation on mRNA
expression results. Therefore, the verification of RNA integrity prior
to downstream applications like RT-qPCR and mircroarrays is
indispensable. A fast and reliable assessment of RNA integrity can be
done with the Eukaryote Total RNA Nano Assay of the Agilent 2100
Bioanalyzer. The importance of RNA quality should also be considered
in new applications such as the investigation of miRNA expression
profiles. With the Agilent Small RNA Assay, Agilent is offering one of
the few possibilities for selectively estimating miRNA before
expression analysis. However, by now little is known about factors
affecting miRNA analysis. Herein, the important impact of total RNA
quality on quantification of mRNA and miRNA should be considered.

RNA quality control in miRNA expression analysis
Agilent Technologies Application Note 5990-5557EN

mRNA and microRNA quality control for RT-qPCR analysis
Methods. 2010 Apr;50(4): 237-243
The importance of high quality sample material, i.e. non-degraded or
fragmented RNA, for classical gene expression profiling is well
documented. Hence, the analysis of RNA quality is a valuable tool in
the preparation of methods like RT-qPCR and microarray analysis. For
verification of RNA integrity, today the use of automated capillary
electrophoresis is state of the art. Following the recently published
MIQE guidelines, these pre-PCR evaluations have to be clearly
documented in scientific publication to increase experimental
transparency. RNA quality control may also be integrated in the
routine analysis of new applications like the investigation of
microRNA (miRNA) expression, as there is little known yet about
factors compromising the miRNA analysis. Agilent Technologies is
offering a new lab-on-chip application for the 2100 Bioanalyzer making
it possible to quantify miRNA in absolute amounts (pg) and as a
percentage of small RNA (%). Recent results showed that this analysis
method is strongly influenced by total RNA integrity. Ongoing RNA
degradation is accompanied by the formation of small RNA fragments
leading to an overestimation of miRNA amount on the chip. Total RNA
integrity is known to affect the performance of RT-qPCR as well as the
quantitative results in mRNA expression profiling. The actual study
identified a comparable effect for miRNA gene expression profiling.
Using a suitable normalization method could partly reduce the
impairing effect of total RNA integrity.

and much more  =>  http://RNA-integrity.gene-quantification.info
- Procedures for Quality Control of RNA Samples for Use in
Quantitative Reverse Transcription PCR.
- Measurable impact of RNA quality on gene expression results from
quantitative PCR.
- Preservation of fine-needle aspiration specimens for future use in
RNA-based molecular testing.
- Maintaining RNA integrity in a homogeneous population of mammary
epithelial cells isolated by Laser Capture Microdissection.
- RNA integrity in post-mortem samples: influencing parameters and
implications on RT-qPCR assays.
- Analysis of Circulating MicroRNA  -  Pre-analytical and Analytical
- Reliable gene expression measurements from degraded RNA by
quantitative real-time PCR depend on short amplicons and a proper
- Evaluation of different RNA extraction methods for small quantities
of plant tissue: Combined effects of reagent type and homogenization
procedure on   RNA quality-integrity and yield.
- Combined expression of miR-122a, miR-1, and miR-200b can
differentiate degraded RNA samples from liver, pancreas, and stomach.
- Maintaining RNA integrity in a homogeneous population of mammary
epithelial cells isolated by Laser Capture Microdissection.
- Robust microRNA stability in degraded RNA preparations from human
tissue and cell samples.
- Assessment of mRNA and microRNA stabilization  in peripheral Human
Blood for Multicenter studies  and Biobanks


Did you miss the qPCR 2011 Symposium in Freising?

BioEPS is now presenting the qPCR 2011 eConference online via video
With our new eConference streaming server a world-wide know-how
transfer becomes possible in a fast and simple way!
qPCR 2011 Event now available NOW as eConfernce =>  http://eConference.qPCR2011.net

At the event around 500 attendee from 47 contries participated from
28th March - 1th April 2011. During the scientific symposium the
presented talks were recorded in excellent sound quality and high
movie resolution.

The qPCR 2011 eConference package contains 65 talks, grouped in
following topics:
Single-Cell qPCR,  Next Generation Sequencing,  High throughput qPCR,
MIQE guidelines,  RNAi-miRNA,  qPCR Data analysis,  Circulating
Nucleic Acids,  Pre-analytical Steps, New qPCR Applications in
Molecular Diagnostics, and more ... ...
qPCR 2011 Event now available NOW as eConfernce =>  http://eConference.qPCR2011.net

You can watch an in-depth demo presentation of our seminars and talks
right here
=>  http://eseminars.bioeps.com/player/?action=demo


MIQE_qPCR APP for iPhone, iPad and iPod  -  iOS Universal

Get help from a special team of experts in qPCR while on the move.
MIQE - qPCR helps you in reviewing scientific works and checking your
own experiments, when qPCR is involved. Check your project's
compliance to MIQE in minutes, have all required references to hand,
and follow qPCR events and news.


Forward Please send the qPCR NEWS to further scientists and friends
who are interested in qPCR !

Best regards,

Michael W. Pfaffl
responsible Editor of the Gene Quantification Pages


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