Quantitation of mRNA by Polymerase Chain Reaction

Nonradioactive PCR Methods

Thomas Köhler, Dirk Laßner, Anne-Katrin Rost, Barbara Thamm, Barbara Pustowoit, Harald Remke (Autoren)

Buch | Softcover

XX, 166 Seiten

2011 | 1995
Springer Berlin (Verlag)
978-3-642-79714-9 (ISBN)

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In this laboratory "cook-book", the authors provide a concise guide to PCR-based techniques to quantify nucleic acids in biological and clinical samples using exclusively nonradioactive detection methods, e.g. HPLC, biotin and digoxigenin based protocols. Each method presentation also includes sections on theory, reagents, standards, applicability, limitations, and trouble shooting. In addition to the protocols, the authors also provide the necessary information on: general aspects of nucleic acid quantitation; design of PCR standards; mRNA purification; cDNA synthesis; solution hybridization; DNA sequencing. This laboratory guide enables professionals as well as beginners to adopt easily quantitative PCR protocols into their own clinical or biomedical research. In this laboratory "cook-book", the authors provide a concise guide to PCR-based techniques for quantifying nucleic acids in biological and clincial samples using exclusively nonradioactive detection methods, e.g. HPLC, biotin and digoxigenin based protocols. Each method presentation also includes sections on theory, reagents, standards, applicability, limitations, and trouble shooting. In addition to the protocols, the authors also provide the necessary information on: general aspects of nucleic acid quantitation; design of PCR standards; mRNA purification; cDNA synthesis; solution hybridization; DNA sequencing. This laboratory guide enables professionals as well as beginners to adopt easily quanatitative PCR protocols into their own clinical or biomedical research.

Thomas Köhler, Prof. Dr. med. Dr. phil. Dipl.-Psych. Dipl.-Math., ist Privatdozent am Psychologischen Institut der Universität Hamburg sowie Dozent an verschiedenen Ausbildungsinstituten für Psychotherapeuten. Er ist Verfasser zahlreicher Monographien, u.a. zu Psychosomatik, Biopsychologie und biologischen Grundlagen psychischer Störungen.

I Theoretical and Methodical Prerequisites for Using PCR to Quantitate Nucleic Acids.- 1.1 General Aspects and Chances of Nucleic Acid Quantitation by PCR.- 1.2 Design of Suitable Primers and Competitor Fragments for Quantitative PCR.- 1.3 Cloning of Short DNA Fragments and In Vitro Transcription to Generate RNA Standards.- 1.4 Direct Non-lsotopic Sequencing of PCR Products or Standards.- 2Conventional Techniques for mRNA Analysis.- 2.1 Isolation of mRNA.- 2.2 Synthesis of cDNA.- 2.3 Qualitative RT-PCR: Amplification of Synthesized mdr-1 cDNA.- 2.4 Single-Tube RT-PCR.- 2.5 Nonradioactive Determination of PCR Products by Using a DIG-Labeled DNA Probe (Dot Blot).- 2.6 Nonradioactive Northern Blot Hybridization with DIG-Labeled DNA Probes.- 3 Semiquantitative and Quantitative Protocols for Measurement of Nucleic Acids by PCR.- 3.1 Quantitation of mRNA by the ELOSA Technique Using External Standards.- 3.2 Semiquantitative Detection of Viral DNA, e.g. for CMV, by Using the DNA Enzyme Immunoassay (DEIA).- 3.3 HPLC-Analysis of Nucleic Acids.- 3.4 Quantitation of Absolute Numbers of mRNA Copies in a cDNA Sample by Competitive PCR.- Acknowledgment.

"...this technical book will be very useful for all customers of PCR. And they are numerous!... Summarizing: an excellent booklet, which we recommend highly and absolutely necessary to all practicians of PCR products and their applications." Cellular and Molecular Biology

Erscheint lt. Verlag	29.12.2011
Reihe/Serie	Springer Lab Manuals
Zusatzinfo	XX, 166 p. 17 illus.
Verlagsort	Berlin
Sprache	englisch
Maße	155 x 235 mm
Gewicht	305 g
Themenwelt	Studium ► 2. Studienabschnitt (Klinik) ► Humangenetik
	Naturwissenschaften ► Biologie ► Biochemie
	Naturwissenschaften ► Chemie ► Analytische Chemie
Schlagworte	Analyse • Biochemical analysis • Biochemische Analyse • Chemical Analysis • Chemische Analyse • DNA • DNA sequencing • enzymes • Hybridization • Klinisch-chemische Analyse • Nucleic acid • Nucleotide • Nukleinsäuren • PCR • Polymerase chain reaction • polymerase Ketten-Reaktion • Quantitative PCR • RNA • sequencing • synthesis • transcription
ISBN-10	3-642-79714-8 / 3642797148
ISBN-13	978-3-642-79714-9 / 9783642797149
Zustand	Neuware