qPCR NEWS - October 2010 - focus on digital PCR

Editor www.Gene-Quantification.info via methods%40net.bio.net (by editor from gene-quantification.info)
Wed Oct 27 10:32:40 EST 2010

qPCR NEWS - October 2010 - focus on digital PCR

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:

NEW - Copy Number Variations  =3D> http://digital-PCR.gene-quantification.i=
qPCR Symposium USA in November 2010
qPCR Application Workshops  -  and more ... ... ...


Digital PCR (dPCR) is a refinement of conventional PCR methods that
can be used to directly quantify and clonally amplify nucleic acids
(including DNA, cDNA, methylated DNA, or RNA). The key difference
between dPCR and traditional PCR lies in the method of measuring
nucleic acids amounts, with the former being a more precise method
than PCR. The main principle of digital PCR is that a single sample is
split into many fractions, all of which are subsequently analysed by a
standard PCR method. The sample is fractionated by the simple process
of dilution so that each fraction contains approximately one copy of
DNA template. This separation allows a more reliable collection and
sensitive measurement of nucleic acid amounts. The other major feature
of digital PCR is that data are recorded as positive or negative (i.e.
generation of an amplification product or not). Hence, the individual
readout signals are qualitative or =91digital=92 in nature. The method has
been demonstrated as useful for studying variations in gene sequences
- such as copy number variants, point mutations, and it is routinely
used for clonal amplification of samples for "next-generation

=3D>  http://digital-PCR.gene-quantification.info


dPCR Reviews and application papers Concept of the limiting dilution

Limiting dilution analysis:  from frequencies to cellular
Dozmorov I, Eisenbraun MD, Lefkovits I.
Immunol Today. 2000 Jan;21(1):15-8.
Dept of Pathology, University of Michigan, Ann Arbor, MI 48109, USA.

End-point limiting-dilution real-time PCR assay for evaluation of
hepatitis C virus quasispecies in serum: performance under optimal and
suboptimal conditions.
Ramachandran S, Xia GL, Ganova-Raeva LM, Nainan OV, Khudyakov Y.
J Virol Methods. 2008 Aug;151(2): 217-224
Division of Viral Hepatitis, Centers for Disease Control and
Prevention, Atlanta, GA 30333, USA

Digital PCR - publications

Digital PCR.
Vogelstein B, Kinzler KW.
Proc Natl Acad Sci U S A. 1999 Aug 3;96(16):9236-41.
The Howard Hughes Medical Institute and the Johns Hopkins Oncology
Center, Baltimore, MD 21231, USA

Nanoliter scale PCR with TaqMan detection.
Kalinina O, Lebedeva I, Brown J, Silver J.
Nucleic Acids Res. 1997 May 15;25(10):1999-2004.
Laboratory of Molecular Microbiology, National Institute of Allergy
and Infectious Diseases, National Institutes of Health, Bethesda MD
20892, USA.

Principle and applications of digital PCR.
Pohl G, Shih IeM.
Expert Rev Mol Diagn. 2004 Jan;4(1): 41-47
Department of Pathology, 418 North Bond Street, B-315, Baltimore, MD
21231, USA.

Microfluidics digital PCR reveals a higher than expected fraction of
fetal DNA in maternal plasma.
Lun FM, Chiu RW, Allen Chan KC, Yeung Leung T, Kin Lau T, Dennis Lo
Clin Chem. 2008 Oct;54(10): 1664-1672.
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing
Institute of Health Sciences, The Chinese University of Hong Kong,
Shatin, New Territories, Hong Kong.

Digital PCR for the molecular detection of fetal chromosomal
Lo YM, Lun FM, Chan KC, Tsui NB, Chong KC, Lau TK, Leung TY, Zee BC,
Cantor CR, Chiu RW.
Proc Natl Acad Sci U S A. 2007 7;104(32): 13116-13121
Li Ka Shing Institute of Health Sciences, Department of Chemical
Pathology, School of Public Health, The Chinese University of Hong
Kong, Sha Tin, New Territories, Hong Kong Special Administrative
Region, People's Republic of China.

Microdissection molecular copy-number counting (microMCC)--unlocking
cancer archives with digital PCR.
McCaughan F, Darai-Ramqvist E, Bankier AT, Konfortov BA, Foster N,
George PJ, Rabbitts TH, Kost-Alimova M, Rabbitts PH, Dear PH.
J Pathol. 2008 216(3): 307-316
Centre for Respiratory Research, Department of Medicine, Royal Free
and University College Medical School, The Rayne Institute, London

Single-molecule genomics.
McCaughan F, Dear PH.
J Pathol. 2010 Jan;220(2):297-306.
MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH,

Microfluidic digital PCR enables multigene analysis of individual
environmental bacteria.
Ottesen EA, Hong JW, Quake SR, Leadbetter JR.
Science. 2006 314(5804): 1464-1467
Division of Biology, California Institute of Technology, Pasadena, CA
91125, USA.

Methylation analysis

DNA methylation analysis by digital bisulfite genomic sequencing and
digital MethyLight.
Weisenberger DJ, Trinh BN, Campan M, Sharma S, Long TI, Ananthnarayan
S, Liang G, Esteva FJ, Hortobagyi GN, McCormick F, Jones PA, Laird PW.
Nucleic Acids Res. 2008 Aug;36(14): 4689-98
Department of Surgery, University of Southern California/Norris
Comprehensive Cancer Center, Los Angeles, CA 90033, USA.

The implications of heterogeneous DNA methylation for the accurate
quantification of methylation
Thomas Mikeska, Ida LM Candiloro, & Alexander Dobrovic
Epigenomics  2010, Vol. 2: 561-573

Rapid analysis of heterogeneously methylated DNA using digital
methylation-sensitive high resolution melting: application to the
CDKN2B (p15) gene.
Candiloro IL, Mikeska T, Hokland P, Dobrovic A.
Epigenetics Chromatin. 2008 Nov 3;1(1): 7
Molecular Pathology Research and Development Laboratory, Department of
Pathology, Peter MacCallum Cancer Centre, Melbourne, Victoria 8006,

GMO analysis

Absolute quantification of genetically modified MON810 maize (Zea mays
L.) by digital polymerase chain reaction.
Corbisier P, Bhat S, Partis L, Xie VR, Emslie KR.
Anal Bioanal Chem. 2010 396(6): 2143-2150

find much more interesting dPCR papers here =3D>  http://digital-PCR.gene-q=


BioEPS GmbH - qPCR Application Workshops

Life Science is still a growing sector and new methods and
technologies are continously developed. Therefore permanent training
and education becomes so important.

With our specific course program we are offering a range of high-
quality course modules, in cooperation with different companies to
give a general and independent overview of existing qPCR technologies
and systems. Our course issues are based on skilled know-how from own
research studies and publications.

Our aim is to point out a critical way of thinking to increase the
quality and outcome of experimental data.

All courses are held regularly in Freising-Weihenstephan, Germany, in
German and English language.
Further customized workshops and specialized trainings will be held as
well across Europe and world-wide.
Workshops are powered by BioEPS GmbH, located at the campus of the
Technical University of Munich, in Freising-Weihenstephan, very close
to the Munich Airport (MUC). For more information and registration,
please see our web page =3D> http://workshops.gene-quantification.info/

Course Occasions 2010:

3-day qPCR Basic Module
2-day BioStatistics & Expression Profiling Module
3-day single-cell qPCR
2-day microRNA qPCR
1-day HRM
2-das qPCR-R data analysis   NEW !
1-day Project Management   NEW !
2-day Quality Management  NEW !

Course dates 2010:

8 - 9  November 2010  (E)   2-day microRNA & qPCR (Mon.-Tue.)
29 November - 1 December 2010  (E)   3-day Experiment Design  &  qPCR
data processing  (Mon. - Wed.)
2 - 3 December 2010  (E)   2-day BioStatistics Module (Thu. - Fri.)

Download course brochure 2010 =3D> http://www.gene-quantification.de/bioeps=

Register here =3D> http://site.bioeps.com/index.php?option=3Dcom_seminar&It=

Access to our workshops =3D> http://www.gene-quantification.de/bioeps-acces=


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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