qPCR NEWS - August 2012 - focus on single-cell qPCR

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qPCR NEWS - August 2012 - focus on single-cell qPCR --------------------------------------------------------------------- Dear researcher, dear Gene Quantification page reader, Our newsletter informs about the latest news in quantitative real-time PCR = (qPCR and RT-qPCR), which are compiled and summarised on the=20 Gene Quantification homepage. The focus of this newsletter issue is: * UPDATE of various new papers using single-cell PCR technologies in qPCR a= nd NGS - http://singlecell.Gene-Quantification.info * Second announcement qPCR & NGS Symposium in March 2013 - http://www.qPCR-= NGS-2013.net=20 * GenEx - a powerful tool for qPCR data analysis - download a free trial ve= rsion - http://GenEx.gene-quantification.info ---------------------------------------------------------------------------= -- If this newsletter is not displayed correctly by your email client, please = use following http://qPCRnews.gene-quantification.info ---------------------------------------------------------------------------= -- UPDATE of various new papers using single-cell PCR technologies in qPCR and= NGS =3D> http://singlecell.Gene-Quantification.info Single-cell molecular-biology is a relatively new scientific branch in biol= ogy. The first single-cell analysis were involved in the characterization o= f mitochondrial DNA in 1988. Single-cell DNA analysis, in particular genomi= c DNA, is important and may be informative in the analysis of genetics of c= ell clonality, genetic anticipation and single-cell DNA polymorphisms. Nowa= days for most sceientists the quantitative transcriptomics in a single-cell= is much more important, and the analytical method of choice is the quantit= ative real-time RT-PCR. In single-cell biology the absolute abundance of pa= rticular mRNAs or microRNAs and their up- or down-regulation in a single ce= ll, compared to their neighbour cells, is the goal. The need for quantitati= ve single-cell mRNA analysis is evident given the vast cellular heterogenei= ty of all tissue cells and the inability of conventional RNA methods, like = northern blotting, RNAse protection assay or classical block RT-PCR, to dis= tinguish individual cellular contributions to mRNA abundance differences.= =20 =20 new papers Method of the year - Methods to watch - Special feature Single-cell methods - Improved single-cell methods are helping to unravel b= iological complexity We present important methods and areas of methodological development worth = watching in the coming years. Nature Methods - VOL.9 NO.1 - JANUARY 2012 - 35 The heterogeneity of cells in culture and in organisms poses a challenge fo= r many experi-mental measurements. Population measure-ments are necessarily= averages, masking the behavior of minority subpopulations and effectively = blinding researchers to possibly interesting differences between cells.The = alternative is to make measure-ments on single cells. Methodologically spea= king, this, too, is challenging on sever-al fronts. Molecular analyses, whe= ther on a particular macromolecule or at an =91omic=92 scale, can be diffic= ult (or even impossible) to accomplish on the amount of mate-rial extracted= from one cell. Methods with increased sensitivity are therefore in demand.= Throughput is also a bottle-neck. Basing firm conclusions on single-cell m= easurements means that one must be able to quickly and accurately analyze m= any cells. Finally, it is often necessary to analyze single cells in a mul-= tiplexed fash-ion, either because the cells exist in a heteroge-neous pop-u= lation or because one wants to measure many parameters at the same time.The= re continue to be methodologi-cal advances on all of these fronts. Mass cyt= ometry, for instance - in which iso-topes are used as antibody labels inste= ad of fluorescent probes=97considerably extends the multiplexing capabiliti= es of flow cytometry (Science 332, 687=96695; 2011). Is the measurement o= f gene expression, digital reverse-transcriptase PCR in a microfluidics de= vice makes it possible to simultaneously monitor the expres-sion of hundred= s of genes in hundreds of single cells. As demonstrated in a recent study o= f tumor heterogeneity, this can be combined with single cell sorting and wi= th statistical clustering methods to begin to dissect the cellular subpopul= ations that constitute a tissue (Nat. Biotechnol. 29, 1120=961127; 2011). ---------------------------------------------------------------------------= ----- Cell Biology. Using cell-to-cell variability - a new era in molecular biolo= gy Pelkmans L. Science. 2012 Apr 27;336(6080): 425-426 ---------------------------------------------------------------------------= ----- Genomic analysis at the single-cell level Kalisky T, Blainey P, Quake SR. Annu Rev Genet. 2011;45: 431-445 Studying complex biological systems such as a developing embryo, a tumor, o= r a microbial ecosystem often involves understanding the behavior and heter= ogeneity of the individual cells that constitute the system and their inter= actions. In this review, we discuss a variety of approaches to single-cell = genomic analysis. ---------------------------------------------------------------------------= ----- A single molecule view of gene expression Larson DR, Singer RH, Zenklusen D. Trends Cell Biol. 2009 (11): 630-637 Analyzing the expression of single genes in single cells appears minimalist= ic in comparison to gene expression studies based on more global approaches= .. However, stimulated by advances in imaging technologies, single-cell stud= ies have become an essential tool in understanding the rules that govern ge= ne expression. This quantitative view of single-cell gene expression is bas= ed on counting mRNAs in single cells, monitoring transcription in real time= , and visualizing single proteins. Parallel advances in mathematical models= based on stochastic, discrete descriptions of biochemical processes have p= rovided crucial insights into the underlying cellular mechanisms that contr= ol expression. The view that has emerged is rooted in a probabilistic under= standing of cellular processes that quantitatively explains both the mean a= nd the variation observed in gene-expression patterns among single cells. T= hus, the close coupling between imaging and mathematical theory has establi= shed single-cell analysis as an essential branch of systems biology. ---------------------------------------------------------------------------= ----- Single-cell gene-expression profiling and its potential diagnostic applicat= ions Stahlberg A, Kubista M, Aman P. Expert Rev Mol Diagn. 2011 (7): 735-740 Gene-expression profiling has been successfully applied in various diagnost= ic applications, but its full capacity is yet to be realized. Samples are g= enerally prepared from a mixture of different cells that are present in unk= nown proportions. Cells are, in many aspects, unique in their characteristi= cs and this heterogeneity confounds the expression profile. The development= of new and robust techniques to measure gene expression in single cells op= ens new avenues in molecular medicine. Today, gene-expression profiles of i= ndividual cells can be measured with high precision and accuracy, identifyi= ng different cell types as well as revealing heterogeneity among cells of t= he same kind. Here, we review practical aspects of single-cell gene-express= ion profiling using reverse transcription quantitative real-time PCR and it= s potential use in diagnostics.=20 ---------------------------------------------------------------------------= -- New papers using single-cell PCR technologies in qPCR and NGS - http://si= nglecell.Gene-Quantification.info - Multimarker gene analysis of circulating tumor cells in pancreatic cancer= patients: a feasibility study - Mammalian genes are transcribed with widely different bursting kinetics - Measuring single-cell gene expression dynamics in bacteria using fluoresc= ence time-lapse microscopy - Quantification noise in single cell experiments - Identifying single-cell molecular programs by stochastic profiling - High-throughput microfluidic single-cell RT-qPCR - RNA-Seq analysis to capture the transcriptome landscape of a single cell - Development and applications of single-cell transcriptome analysis - Quantitative RT-PCR gene expression analysis of laser microdissected tiss= ue samples - mRNA and microRNA expression profiles in circulating tumor cells and prim= ary tumors of metastatic breast cancer patients - Comprehensive qPCR profiling of gene expression in single neuronal cells - Single cell transcriptomics of neighboring hyphae of Aspergillus niger - RT-qPCR based quantitative analysis of gene expression in single bacteria= l cells - Circulating tumor cells in breast cancer: detection systems, molecular ch= aracterization, and future challenges - Molecular characterization of circulating tumor cells in breast cancer: c= hallenges and promises for individualized cancer treatment - Gene expression profile of circulating tumor cells in breast cancer by RT= -qPCR - Single-cell gene-expression profiling reveals qualitatively distinct CD8 = T cells elicited by different gene-based vaccines - High throughput single cell expression profiling: Taking a closerlook on = biological response - Quantification of circulating endothelial and progenitor cells: compariso= n of quantitative PCR and four-channel flow cytometry - Parthenogenic blastocysts derived from cumulus-free in vitro matured huma= n oocytes ---------------------------------------------------------------------------= -- Second announcement qPCR & NGS Symposium in Freising-Weihenstephan 18-22 March 2013 http://www.qPCR-NGS-2013.net Download the latest Event Flyer =3D> http://tinyurl.com/qPCR-NGS-2013 On behalf of the Organisation Committee and the Scientific Board it is a gr= eat pleasure to invite you to the 6th International qPCR & NGS 2013 Event. = The event is divided in a 3-day scientific Symposium with an Industrial Exh= ibition and various 2-day Application Workshop to be held at the Center of = Life Science in Freising Weihenstephan, Technische Universit=E4t M=FCnchen = (Germany). The great international interest in the previous meetings ( qPCR= 2004 to qPCR 2011 ) led us to the decision to repeat the Symposium in spri= ng 2013. We expect 500-600 participants coming from all over the world, in = 2011 we could welcome participants from 56 contries, and roughly 30-40 inte= rnational companies in the qPCR Industrial Exhibition. We have set the date for the qPCR & NGS 2013 Event to 18th - 22nd March 201= 3. The event location is the central lecture hall complex and the foyer at = TUM (Technical University of Munich) in Freising Weihenstephan, Germany (Go= ogle Maps link or Google Earth link). The TUM and the Biotech region around= Munich is part of the largest Biotech cluster in Europe, located close to = the Munich airport in the heart of Bavaria. The focus of the qPCR & NGS 2013 Event will be on: Next Generation Thinki= ng in Molecular Diagnostics Leading academic researchers and industrial contributors in the field will = participate in the symposium, which will be an arena for fruitful discussio= ns between researchers of different backgrounds. The Symposium Talks, Poste= r Sessions, Industrial Exhibition and associated qPCR & NGS Application Wor= kshops offer an overview of the present knowledge and future developments i= n qPCR, next generation sequencing and gene expression measurement technolo= gy and its wide applications in research. The symposium will focus on 70 lectures and a huge poster exhibition will b= e presented by internationally recognised experts in their field. The empha= sis will be on unbiased, didactic information exchange. Internationally ren= own speakers will be participating in a lively and exciting programme enabl= ing the valuable exchange of information in the qPCR and Next Generation Se= quencing field. One third of the talks will be presented by selected invite= d speakers, one third will be selected from the submitted abstracts and one= third will be presented by qPCR & NGS related company R&D representatives.= All scientific contributions will be published in the Symposium Proceeding= s (ISBN to be announced). Full papers from selected invited academic and industrial speakers and appl= ication notes from industrial speakers will be published in a METHODS speci= al issue =93Transcriptional Biomarkers=94 edited by Michael W. Pfaffl (pub= lished January 2013). At the meeting all participants will get a free har= d cover of this special Methods issue. Please have a look to our previous i= ssue =3D> =93The ongoing evolution of qPCR=94 METHODS special qPCR Vol 50 = issue 4 (April 2010) Please register using the Internet based ConfTool registration and submissi= on platform =3D> http://registration.qPCR-NGS-2013.net ---------------------------------------------------------------------------= -- Symposium Talk and Poster sessions: http://sessions.qpcr-ngs-2013.net/ - Main Topic: Molecular diagnostics - Main Topic: Next Generation Sequencing (NGS) - Main Topic: Transcriptional Biomarkers - High throughput analysis in qPCR - Systems biology - Single-cells diagnostics - MIQE & QM strategies in qPCR - non-coding RNAs - microRNA, siRNA, long non-coding RNAs - Digital PCR & Nano-fluidics - Pre-analytical Steps - BioStatistics & BioInformatics - qPCR & NGS data analysis - Lunch Seminars: - qBASEplus - data analysis lunch seminar - GenEx - data analysis lunch seminar - NGS data analysis lunch seminars - more to be announced........ View our qPCR 2011 event trailer on YouTube =3D> http://www.youtube.com/wat= ch?v=3Dcp8WwPyLW8Y Download the latest Event Flyer =3D> http://tinyurl.com/qPCR-NGS-2013 Please register using the Internet based ConfTool registration and submissi= on platform =3D> http://registration.qPCR-NGS-2013.net ---------------------------------------------------------------------------= -- GenEx 5 - A Powerful Tool For qPCR Data Analysis Download a free trail version here =3D> http://GenEx.gene-quantification.in= fo GenEx is a popular software for qPCR data processing and analysis. Built in= a modular fashion GenEx provides a multitude of functionalities for the qP= CR community, ranging from basic data editing and management to advanced cu= tting-edge data analysis. View our webpage =3D> http://GenEx.gene-quantific= ation.info Basic data editing and management Arguably the most important part of qPCR experiments is to pre-process the = raw data into shape for subsequent statistical analyses. The pre-processing= steps need to be performed consistently in correct order and with confiden= ce. GenEx Standard=92s streamlined and user-friendly interface ensures mist= ake-free data handling. Intuitive and powerful presentation tools allow pro= fessional illustrations of even the most complex experimental designs. Advanced cutting-edge data analysis When you need more advanced analyses GenEx Enterprise is the product for yo= u. Powerful enough to demonstrate feasibility it often proves sufficient fo= r most users demands. Current features include parametric and non-parametri= c statistical tests, Principal Component Analysis, and Artificial Neural Ne= tworks. New features are continuously added to GenEx with close attention t= o customers=92 needs. New features Sample handling and samples individual biology often contribute to confound= ing experimental variability. By using the new nested ANOVA feature in GenE= x version 5 user will be able to evaluate variance contributions from each = step in the experimental procedure. With a good knowledge of the variance c= ontributions, an appropriate distribution of experimental replicates can be= selected to minimize confounding variance and maximize the power of the ex= perimental design! For experiments with complex features, such as for examp= le multifactorial diseases, analytical relationships and classifications ma= y not readily be available. The support vector machine feature in the new v= ersion of GenEx is so easy to use that it will make this advanced supervise= d classification method easily available to novice users, while providing a= ccess to advanced parameters for experts. Download a free trail version here =3D> http://GenEx.gene-quantification.in= fo GenEx PDF user guides: * GenEx user guide=20 * GenEx user guide - Exiqon Wizard=20 * GenEx user guide - Roche Wizard ---------------------------------------------------------------------------= -- Please forward this qPCR NEWS http://api.addthis.com/oexchange/0.8/forward= /email/offer?url=3Dhttp://qPCRnews.gene-quantification.info&title=3DJoin+ou= r+monthly+newsletter+on&username=3DqPCR-NEWS&email_template=3D&lng=3Den-us = to further scientists and friends who are interested in qPCR ! Best regards, Michael W. Pfaffl responsible Editor of the Gene Quantification Pages If this newsletter is not displayed correctly by your email client, please = use following=20 LINK http://qPCRnews.gene-quantification.info/ ---------------------------------------------------------------------------= -- The qPCR NEWS and the Gene Quantification Pages are educational sites with = the only purpose of facilitating access to qPCR related information on the = internet. The qPCR NEWS and the Gene Quantification Pages are edited by Mic= hael W. Pfaffl. Copyright =A92005-2012 All rights reserved. Any unauthorized use, reproduct= ion, or transfer of this message or its contents, in any medium, is strictl= y prohibited. 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