RT-PCR Protocols

Part I. Introduction

RT-PCR in Biomedicine: Opportunities Arising from the New Accessibility of mRNA
Joe O'Connell

The Basics of RT-PCR: Some Practical Considerations
Joe O'Connell

Part II. Highly Sensitive Detection and Analysis of mRNA

Description of Quantitative Competitive RT-PCR Technique to Analyze Minute Amounts of Different mRNAs in Small Tissue Samples
Susanne Greber-Platzer, Brigitte Balcz, Christine Fleischmann, and Gert Lubec

Detection of mRNA Expression and Alternative Splicing in a Single Cell
Tsutomu Kumazaki

Nested RT-PCR: Sensitivity Controls are Essential to Determine the Biological Significance of Detected mRNA
Triona Goode, Wen-Zhe Ho, Terry O'Connor, Sandra Busteed, Steven D. Douglas, Fergus Shanahan, and Joe O'Connell

Part III. Quantitative RT-PCR

Quantitative RT-PCR: A Review of Current Methodologies
Caroline Joyce

Rapid Development of a Quantitative-Competitive (QC) RT-PCR Assay Using a Composite Primer Approach
Joe O'Connell, Aileen Houston, Raymond Kelly, Darren O'Brien, Aideen Ryan, Michael W. Bennett, and Kenneth Nally

Quantitation of Gene Expression by RT-PCR and HPLC Analysis of PCR Products
Franz Bachmair, Christian G. Huber, and Guenter Daxenbichler

Time-Resolved Fluorometric Detection of Cytokine mRNAs Amplified by RT-PCR
Kaisa Nieminen, Markus Halminen, Matti Waris, Mika Mäkelä, Johannes Savolainen, Minna Sjöroos, and Jorma Ilonen

Mimic-Based RT-PCR Quantitation of Substance P mRNA in Human Mononuclear Phagocytes and Lymphocytes
Jian-Ping Lai, Steven D. Douglas, and Wen-Zhe Ho

Part IV. Detection and Analysis of RNA Viruses

Detection and Quantification of the Hepatitis C Viral Genome
Liam J. Fanning

Semi-Quantitative Detection of Hepatitis C Virus RNA by "Real-Time" RT-PCR
Joerg F. Schlaak

RT-PCR for the Assessment of Genetically Heterogenous Populations of the Hepatitis C Virus
Brian Mullan, Liam Fanning, Fergus Shanahan, and Daniel G. Sullivan

Part V. In Situ Localization of mRNA Expression

In Situ Immuno-PCR: A Newly Developed Method for Highly Sensitive Antigen Detection In Situ
Yi Cao

RT-PCR from Laser Capture Microdissected Samples
Tatjana Crnogorac-Jurcevic, Torsten O. Nielsen, and Nick R. Lemoine

Mycobacterium Paratuberculosis Detected by Nested PCR in Intestinal Granulomas Isolated by Laser Capture Microdissection in Cases of Crohn's Disease
Paul Ryan, Simon Aarons, Michael W. Bennett, Gary Lee, Gerald C. O'Sullivan, Joe O'Connell, and Fergus Shanahan

RT-PCR-Based Approaches to Generate Probes for mRNA Detection by In Situ Hybridization
Joe O'Connell

Part VI. Differential mRNA Expression

Amplified RNA for Gene Array Hybridizations
Valentina I. Shustova and Stephen J. Meltzer

Semiquantitative Determination of Differential Gene Expression in Primary Tumors and Matched Metastases by RT-PCR: Comparison with Other Methods
Benno Mann and Christoph Hanski

Part VII. Genetic Analysis

Detection of Single Nucleotide Polymorphisms Using a Non-Isotopic RNase Cleavage Assay
Frank Waldron-Lynch, Claire Adams, Michael G. Molloy, and Fergal O'Gara

Part VIII. RT-PCR in Immunology

Detection of Clonally Expanded T Cells by RT-PCR-SSCP and Nucleotide Sequencing of T Cell Receptor b-CDR3 Regions
Manae Suzuki Kurokawa, Kusuki Nishioka, and Tomohiro Kato

Generation of scFv from a Phage Display Mini-Library Derived from Tumor-Infiltrating B Cells
Nadège Gruel, Beatrix Kotlan, Marie Beuzard, and Jean-Luc Teillaud

Generation of Murine scFv Intrabodies from B Cell Hybridomas
Chang Hoon Nam, Sandrine Moutel, and Jean-Luc Teillaud

Quantitation of mRNA Levels by RT-PCR in Cells Purified by FACS: Application to Peripheral Cannabinoid Receptors in Leukocyte Subsets
Jean Marchand and Pierre Carayon

Part IX. RT-PCR in Antisense Technology

Detection of Antisense RNA Transcripts by Antisense RT-PCR
Michael C. Yeung and Allan S. Lau

Part X. RT-PCR in cDNA Cloning

RT-PCR in cDNA Library Construction
Vincent Healy

An RT-PCR-Based Protocol for the Rapid Generation of Large, Representative cDNA Libraries Suitable for Expression Screening
Joe O'Connell

Index