PCR CLONING PROTOCOLS
CONTENTS PART I PERFORMING AND OPTIMIZING PCR 1 POLYMERASE CHAIN REACTION: BASIC PRINCIPLES AND ROUTINE PRACTICE 2 COMPUTER PROGRAMS FOR PCR PRIMER DESIGN AND ANALYSIS 3 SINGLE-STEP PCR OPTIMIZATION USING TOUCHDOWN AND STEPDOWN PCR PROGRAMMING 4 XL PCR AMPLIFICATION OF LONG TARGETS FROM GENOMIC DNA 5 COUPLED ONE-STEP REVERSE TRANSCRIPTION AND POLYMERASE CHAIN REACTION PROCEDURE FOR CLONING LARGE CDNA FRAGMENTS 6 LONG DISTANCE REVERSE-TRANSCRIPTION PCR 7 INCREASING PCR SENSITIVITY FOR AMPLIFICATION FROM PARAFFIN- EMBEDDED TISSUES 8 GC-RICH TEMPLATE AMPLIFICATION BY INVERSE PCR: DNA POLYMERASE AND SOLVENT EFFECTS 9 PCR PROCEDURE FOR THE ISOLATION OF TRINUCLEOTIDE REPEATS 10 METHYLATION-SPECIFIC PCR 11 DIRECT CLONING OF FULL-LENGTH CELL DIFFERENTIALLY EXPRESSED GENES BY MULTIPLE ROUNDS OF SUBTRACTIVE HYBRIDIZATION BASED ON LONG-DISTANCE PCR AND MAGNETIC BEADS PART II CLONING PCR PRODUCTS 12 CLONING PCR PRODUCTS: AN OVERVIEW 13 USING T4 DNA POLYMERASE TO GENERATE CLONABLE PCR PRODUCTS 14 ENZYME-FREE CLONING OF PCR PRODUCTS AND FUSION PROTEIN EXPRESSION 15 DIRECTIONAL RESTRICTION SITE-FREE INSERTION OF PCR PRODUCTS INTO VECTORS 16 AUTOSTICKY PCR: DIRECTIONAL CLONING OF PCR PRODUCTS WITH PREFORMED 5' OVERHANGS 17 A RAPID AND SIMPLE PROCEDURE FOR DIRECT CLONING OF PCR PRODUCTS INTO BACULOVIRUSES PART III MUTAGENESIS AND RECOMBINATION 18 PCR APPROACHES TO DNA MUTAGENESIS AND RECOMBINATION: AN OVERVIEW 19 IN-FRAME CLONING OF SYNTHETIC GENES USING PCR INSERTS 20 MEGAPRIMER PCR 21 PCR-MEDIATED RECOMBINATION: A GENERAL METHOD APPLIED TO CONSTRUCT CHIMERIC INFECTIOUS MOLECULAR CLONES 22 PCR METHOD FOR GENERATING MULTIPLE MUTATIONS AT ADJACENT SITES 23 A FAST POLYMERASE CHAIN REACTION-MEDIATED STRATEGY FOR INTRODUCING REPEAT EXPANSIONS INTO CAG-REPEAT CONTAINING GENES 24 PCR SCREENING IN SIGNATURE-TAGGED MUTAGENESIS OF ESSENTIAL GENES 25 STAGGERED EXTENSION PROCESS (STEP) IN VITRO RECOMBINATION 26 RANDOM MUTAGENESIS BY WHOLE-PLASMID PCR AMPLIFICATION PART IV CLONING UNKNOWN NEIGHBORING DNA 27 PCR-BASED STRATEGIES TO CLONE UNKNOWN DNA REGIONS FROM KNOWN FOREIGN INTEGRANTS: AN OVERVIEW 28 LONG DISTANCE VECTORETTE PCR (LDV PCR) 29 NONSPECIFIC, NESTED SUPPRESSION PCR METHOD FOR ISOLATION OF UNKNOWN FLANKING DNA (“COLD-START METHOD”) 30 INVERSE PCR: CDNA CLONING 31 INVERSE PCR: GENOMIC DNA CLONING 32 GENE CLONING AND EXPRESSION PROFILING BY RAPID AMPLIFICATION OF GENE INSERTS WITH UNIVERSAL VECTOR PRIMERS 33 THE ISOLATION OF DNA SEQUENCES FLANKING TN5 TRANSPOSON INSERTIONS BY INVERSE PCR 34 RAPID AMPLIFICATION OF GENOMIC DNA SEQUENCES TAGGED BY INSERTIONAL MUTAGENESIS 35 ISOLATION OF LARGE TERMINAL SEQUENCES OF BAC INSERTS BASED ON DOUBLE-RESTRICTION-ENZYME DIGESTION FOLLOWED BY ANCHORED PCR 36 A “STEP DOWN” PCR-BASED TECHNIQUE FOR WALKING INTO AND THE SUBSEQUENT DIRECT SEQUENCE ANALYSIS OF FLANKING GENOMIC DNA PART V LIBRARY CONSTRUCTION AND SCREENING 37 USE OF PCR IN LIBRARY SCREENING: AN OVERVIEW 38 CLONING OF HOMOLOGOUS GENES BY GENE-CAPTURE PCR 39 RAPID AND NONRADIOACTIVE SCREENING OF RECOMBINANT LIBRARIES BY PCR 40 RAPID CDNA CLONING BY PCR SCREENING (RC-PCR) 41 GENERATION AND PCR SCREENING OF BACTERIOPHAGE ? SUBLIBRARIES ENRICHED FOR RARE CLONES (THE “SUBLIBRARY METHOD”) 42 PCR-BASED SCREENING FOR BACTERIAL ARTIFICIAL CHROMOSOME LIBRARIES 43 A 384-WELL MICROTITER-PLATE-BASED TEMPLATE PREPARATION AND SEQUENCING METHOD 44 A MICROTITER-PLATE-BASED HIGH THROUGHPUT PCR PRODUCT PURIFICATION METHOD