PCR has developed to the most powerful tool for the study of DNA and genes. Various techniques based on PCR are described in this manual, including protocols for the analysis of genes, mutations, abnormal gene expression in tumors and gene expression during differentiation.

Klappentext

DNA and RNA fingerprinting based on arbitrarily primed PCR provides the most powerful tool for the study of genes.
The basic techniques are described in detailed protocols including each step from template preparation to fingerprint visualization. Various protocols for the basic techniques allow to choose between alternative strategies. In addition to the general techniques specific research applications of particular interest are given such as gene mapping, detection of somatic mutations, gene abnormally expressed in tumors or differentially expressed genes by RNA fingerprinting.

Inhalt

Overview.- I DNA Extraction from Mammals.- II Insect DNA Extraction Protocol.- III Rapid DNA Extraction from Plants.- IV Preparation of Fungal Genomic DNA for PCR and RAPD Analysis.- V Extraction of Histoplasma capsulatum DNA for PCR.- VI DNA Extraction from Bacterial Cultures.- VII Arbitrarily Primed PCR and RAPDs.- VIII Random Amplified Polymorphic DNA Assay.- IX DNA Amplification Fingerprinting.- X Fluorescent Detection and Analysis of RAPD Amplicons Using the ABI PRISM DNA Sequencers.- XI Optimization of RAPD Fingerprinting.- XII Fingerprint Tailoring.- XIII Resolving DNA Amplification Products Using Polyacrylamide Gel Electrophoresis and Silver Staining.- XIV Denaturing Gradient Gel Electrophoresis for Enhanced Detection of DNA Polymorphisms.- XV Modified Temperature Sweep Gel Electrophoresis for the Separation of Arbitrarily Amplified DNA Fragments.- XVI High Throughput Scoring of RAPD Fragments Through the Use of Dot-Blot Hybridization.- XVII Recovering Amplified DNA from Silver Stained Gels.- XVIII Cloning of RAPD Markers.- XIX Sequencing of RAPD Markers.- XX Analysis of Tumor-Specific Genetic Alterations by Arbitrarily Primed PCR.- XXI Construction of Linkage Maps with RAPD Markers.- XXII Pseudo-Testcross Mapping Strategy Using RAPD Markers.- XXIII Estimating Nucleotide Divergence with RAPD Data.- XXIV RAPD and PAUP Analysis for Microbial Screening Programs.- XXV Production of Specific Probes for Microorganisms.- Overview.- XXVI Differential Display of Expressed mRNAs.- XXVII RNA Arbitrarily Primed PCR.- XXVIII Nonradioactive Differential Display of Messenger RNA.- XXIX Fluorescent Differential Display: A Fast and Safe Way for Reliable Differential Display Analysis.- XXX Slot Blot Hybridization Screening.- XXXI How To Find and Clone the Appropriate cDNA Fragments Generated in Differential mRNA Display by Using Northern Blot for cDNA Capture.- XXXII Verification of Differential Display Results by RNase Protection.- XXXIII Direct Automated Sequencing of DDRT-PCR Fragments.- XXXIV Ligation Linked PCR and Direct Sequencing of Differential Display Products.- XXXV Differential Display PCR Fragments as Probes for cDNA Cloning.- XXXVI Targeted RNA Fingerprinting.- XXXVII Generation of Tumor-Specific DEST Catalogue.- XXXVIII Analysis of Gene Expression in the Preimplantation Mouse Embryo Using mRNA Differential Display.- XXXIX Rapid Identification of Differentially Expressed Genes in Trypanosomes.