Parasite Genomics Protocols

Parasitic disease remains an important health issue in most regions of the world and constitutes a major cause of morbidity and mortality in developing countries. In Parasite Genomics Protocols, pioneering bench researchers describe in detail the cutting-edge techniques they have developed for analyzing the genomes and gene products in a diverse range of protozoan and metazoan parasites. These readily reproducible techniques can be used in genomic, functional genomic, and postgenomic studies and include transfection methods and vectors for several protozoan parasites, global analysis using microarrays, gene ablation using RNA interference, gene knockout, mutagenesis, and chromosome manipulation. Some of the protocols require DNA sequence data, whereas others were developed independently of whole-genome sequence data. Additional chapters survey the available genome sequences, helping novice researchers overcome the difficulties of accessing the databases and applying bioinformatics analysis to biological function. The protocols presented follow the successful Methods in Molecular BiologyTM series format, each one offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of equipment and reagents, and tips on troubleshooting and the avoidance of known pitfalls. State-of-the-art and highly practical, Parasite Genomics Protocols offers bench investigators in parasitology a wide-ranging collection of diverse methods for elucidating the content of the genomes of parasitic organisms, utilizing the genome sequence for genetic analysis, and ascribing function to the available genome sequences. Sequencing Strategies For Parasite Genomes / Daniella Bartholomeu And Najib M. El-sayed -- Annotation Of Parasite Genomes / Matthew Berriman And Midori Harris -- Parasite Genome Databases And Web-based Resources / Christiane Hertz-fowler And Neil Hall -- Expressed Sequence Tags: Medium Throughput Protocols / Claire Whitton, Jennifer Daub, Marian Thompson, And Mark Blaxter -- Expressed Sequence Tags: Analysis And Annotation / John Parkinson And Mark Blaxter -- Race And Rage Cloning In Parasitic Microbial Eukaryotes / Bryony A. P. Williams And Robert P. Hirt. Amplified (restriction) Fragment Length Polymorphism (aflp) Analysis / Daniel K. Masiga And C. Michael R. Turner -- Minisatellites And Mvr-pcr For The Individual Identification Of Parasite Isolates / Annette Macleod -- Analysis Of Differentially Expressed Parasite Genes And Proteins Using Transcriptomics And Proteomics / Daniel C. Gare -- Gene Expression Studies Using Self-fabricated Parasite Cdna Microarrays / Karl F. Hoffmann And Jennifer M. Fitzpatrick -- Dna Content Analysis On Microarrays / Upinder Singh, Preetam H. Shah, And Ryan C. Macfarlane. Typing Single Nucleotide Polymorphisms In Toxoplasma Gondii By Allele-specific Primer Extension And Microarray Detection / Chunlei Su, Christian Hott, Bernard H. Brownstein, And L. David Sibley -- Transfection Of The Human Malaria Parasite Plasmodium Falciparum / Brendan S. Crabb, Melanie Rug, Tim-wolf Gilberger, Jennifer K. Thompson, Tony Triglia, Alexander G. Maier, And Alan F. Cowman -- A Pcr-based Method For Gene Deletion And Protein Tagging In Trypanosoma Brucei / George K. Arhin, Shuiyuan Shen, Elisabetta Ullu, And Christian Tschudi. Analysis Of Gene Function In Trypanosoma Brucei Using Rna Interference / Appolinaire Djikeng, Shuiyuan Shen, Christian Tschudi, And Elisabetta Ullu -- In Vitro Shuttle Mutagenesis Using Engineered Mariner Transposons / Kelly A. Robinson, Sophie Goyard, And Stephen M. Beverley -- Random Mutagenesis Strategies For Construction Of Large And Diverse Clone Libraries Of Mutated Dna Fragments / Sudsanguan Chusacultanachai And Yongyuth Yuthavong -- Separation, Digestion And Cloning Of Intact Parasite Chromosomes Embedded In Agarose / Vanessa Leech, Michael A. Quail, And Sara E. Melville. Chromosome Fragmentation In Leishmania / Pascal Dubessay, Christine Blaineau, Patrick Bastien, And Michel Pagès -- Fish Mapping For Helminth Genomes / Hirohisa Hirai And Yuriko Hirai -- Fiber-fish: Fluorescence In Situ Hybridization On Stretched Dna / Klaus Ersfeld -- Yeast Two-hybrid Assay For Studying Protein-protein Interactions / Ahmed Osman -- From Genomes To Vaccines For Leishmaniasis / Carmel B. Stober. Edited By Sara E. Melville. Includes Bibliographical References And Index. Sequencing strategies for parasite genomes / Daniella Bartholomeu and Najib M. El-Sayed Annotation of parasite genomes / Matthew Berriman and Midori Harris Parasite genome databases and web-based resources / Christiane Hertz-Fowler and Neil Hall Expressed sequence tags: medium throughput protocols / Claire Whitton, Jennifer Daub, Marian Thompson, and Mark Blaxter Expressed sequence tags: analysis and annotation / John Parkinson and Mark Blaxter Positive selection scanning of parasite DNA sequences Winston A. Hide and Raphael D. Isokpehi Race and rage cloning in parasitic microbial eukaryotes / Bryony A. P. Williams and Robert P. Hirt Amplified (restriction) fragment length polymorphism (AFLP) analysis / Daniel K. Masiga and C. Michael R. Turner Minisatellites and MVR-PCR for the individual identification of parasite isolates / Annette MacLeod Analysis of differentially expressed parasite genes and proteins using transcriptomics and proteomics / Daniel C. Gare Gene expression studies using self-fabricated parasite cDNA microarrays / Karl F. Hoffmann and Jennifer M. Fitzpatrick DNA content analysis on microarrays / Upinder Singh, Preetam H. Shah, and Ryan C. MacFarlane Typing single nucleotide polymorphisms in toxoplasma gondii by allele-specific primer extension and microarray detection / Chunlei Su, Christian Hott, Bernard H. Brownstein, and L. David Sibley Transfection of the human malaria parasite plasmodium falciparum / Brendan S. Crabb, Melanie Rug, Tim-Wolf Gilberger, Jennifer K. Thompson, Tony Triglia, Alexander G. Maier, and Alan F. Cowman A PCR-based method for gene deletion and protein tagging in trypanosoma brucei / George K. Arhin, Shuiyuan Shen, Elisabetta Ullu, and Christian Tschudi Analysis of gene function in trypanosoma brucei using RNA interference / Appolinaire Djikeng, Shuiyuan Shen, Christian Tschudi, and Elisabetta Ullu In vitro shuttle mutagenesis using engineered mariner transposons / Kelly A. Robinson, Sophie Goyard, and Stephen M. Beverley Random mutagenesis strategies for construction of large and diverse clone libraries of mutated DNA fragments / Sudsanguan Chusacultanachai and Yongyuth Yuthavong Separation, digestion and cloning of intact parasite chromosomes embedded in agarose / Vanessa Leech, Michael A. Quail, and Sara E. Melville Chromosome fragmentation in leishmania / Pascal Dubessay, Christine Blaineau, Patrick Bastien, and Michel PagÅs FISH mapping for helminth genomes / Hirohisa Hirai and Yuriko Hirai Fiber-FISH: fluorescence in situ hybridization on stretched DNA / Klaus Ersfeld Yeast two-hybrid assay for studying protein-protein interactions / Ahmed Osman From genomes to vaccines for leishmaniasis / Carmel B. Stober This volume describes the status of knowledge on the genomes of parasites causing major parasitic diseases, the methods used to analyze the genomes, and how and where to access the data. It details the most recent methods being developed for functional genomic analyses Front Matter....Pages i-xiv Back Matter....Pages 1-16 ....Pages 17-43