Includes supplementary material: sn.pub/extras
Genomic imprinting is the process by which gene activity is regulated according to parent of origin. Usually, this means that either the maternally inherited or the paternally inherited allele of a gene is expressed while the opposite allele is repressed. The phenomenon is largely restricted to mammals and flowering plants and was first recognized at the level of whole genomes. Nuclear transplantation experiments carried out in mice in the late 1970s established the non-equivalence of the maternal and paternal genomes in mammals, and a similar conclusion was drawn from studies of interploidy crosses of flowering plants that extend back to at least the 1930s. Further mouse genetic studies, involving animals carrying balanced translocations (reviewed in Chapter 3), indicated that imprinted genes were likely to be widely scattered and would form a minority within the mammalian genome. The first imprinted genes were identified in the early 1990s; over forty are now known in mammals and the list continues steadily to expand.
Generation of Monoparental Embryos for Investigation into Genomic Imprinting Wendy L. Dean, Gavin Kelsey, and Wolf Reik Deriving and Propagating Mouse Embryonic Stem Cell Lines for Studying Genomic Imprinting Jeffrey R. Mann Balanced Translocations for the Analysis of Imprinted Regions of the Mouse Genome Anne C. Ferguson-Smith, Maxine Tevendale, Pantelis Georgiades, and Valerie Grandjean Production of YAC Transgenic Mice by Pronuclear Injection Justin F.-X. Ainscough, Rosalind M. John, and Sheila C. Barton A Transgenic Approach to Studying Imprinted Genes: Modified BACs and PACs Rosalind M. John, Justin F.-X. Ainscough, and Sheila C. Barton Methylation-Sensitive Genome Scanning Izuho Hatada and Tsunehiro Mukai Subtraction-Hybridization Method for the Identification of Imprinted Genes Fumitoshi Ishino, Yoshimi Kuroiwa, Naoki Miyoshi, Shin Kobayashi, Takashi Kohda, and Tomoko Kaneko-Ishino Identification of Imprinted Loci by Methylation: Use of Methylation- Sensitive Representational Difference Analysis (Me-RDA) Rachel J. Smith and Gavin Kelsey Ribonuclease Protection Joanne L. Thorvaldsen and Marisa S. Bartolomei Quantitative RT-PCR-Based Analysis of Allele-Specific Gene Expression Judith Singer-Sam and Chunguang Gao Allele-Specific In Situ Hybridization (ASISH) Rolf Ohlsson, Kristian Svensson, Hengmi Cui, Helena Malmikumpu, and Gail Adam RNA-FISH to Analyze Allele-Specific Expression Giovanna Braidotti Flow Cytometry and FISH to Investigate Allele-Specific Replication Timing and Homologous Association of Imprinted Chromosomes Janine LaSalle and Marc Lalande Southern Analysis Using Methyl-Sensitive Restriction Enzymes Tom Moore A PCR-Based Method for Studying DNA Methylation Mira Ariel Bisulfite-Based Methylation Analysis of Imprinted Genes Sabine Engemann, Osman El-Maarri, Petra Hajkova, Joachim Oswald, and Joern Walter Direct Analysis of Chromosome Methylation Déborah Bourc'his and Evani Viegas-Péquignot In Vitro Methylation of Predetermined Regions in Recombinant DNA Constructs Ilse Van den Wyngaert, Roger L. P. Adams, and Stefan U. Kass In Vitro Methylation of Specific Regions in Recombinant DNA Constructs by Excision and Religation Ghislaine Dell, Marika Charalambous, and Andrew Ward Detection of Methyl-Sensitive DNA-Binding Proteins with Possible Involvement in the Imprinting Phenomenon Kerstin Otte and Björn Rozell Probing Chromatin Structure with Nuclease Sensitivity Assays Richard I. Gregory, Sanjeev Khosla, and Robert Feil Examining Histone Acetylation at Specific Genomic Regions Ji-Fan Hu and Andrew R. Hoffman Purification of the MeCP2/Histone Deacetylase Complex from Xenopus laevis Peter L. Jones, Paul A. Wade, and Alan P. Wolffe Reconstitution of Chromatin In Vitro Kiyoe Ura and Yasufumi Kaneda Genomic Imprinting in Plants Rinke Vinkenoog, Melissa Spielman, Sally Adams, Hugh G. Dickinson, and Rod J. Scott
Imprinted genes, many of which generally control growth and development, frequently lose their imprints during cancer progression, a loss that then plays a substantial role in uncontrolled tumor growth. Imprint instability also appears to be a major limitation to the success of mammalian cloning experiments. In Genomic Imprinting: Methods and Protocols, Andrew Ward and a team of experienced researchers have brought together a collection of optimized classic and vanguard techniques for the identification and analysis of imprinted genes. The majority of protocols describe molecular techniques that allow examination of gene structure or expression in an allele-specific manner. Protocols are included for identifying and cloning imprinted genes, for analyzing imprinted gene expression, for the study of DNA methylation and methylation-sensitive DNA-binding proteins, and for examining chromatin structure. There are also methods for the manipulation of mouse embryos to produce monoparental embryos and embryonic stem cells, and for the generation of transgenic mice with BAC, PAC, and YAC constructs. Each technique is described in step-by-step detail to ensure successful results.
Incorporating a wealth of knowledge from leading exponents in the field, Genomic Imprinting: Methods and Protocols brings together all the essential molecular, genetic, and embryological methods commonly used in today's laboratories for the identification and analysis of imprinted genes.
"...a multi-authored volume in which the methods of gene imprinting are discussed in detail in 25 chapters. The protocols are very detailed with multiple schemes and figures." - Journal of Pediatric Endocrinology and Metabolism
Generation of Monoparental Embryos for Investigation into Genomic Imprinting.- Deriving and Propagating Mouse Embryonic Stem Cell Lines for Studying Genomic Imprinting.- Balanced Translocations for the Analysis of Imprinted Regions of the Mouse Genome.- Production of YAC Transgenic Mice by Pronuclear Injection.- A Transgenic Approach to Studying Imprinted Genes.- Methylation-Sensitive Genome Scanning.- Subtraction-Hybridization Method for the Identification of Imprinted Genes.- Identification of Imprinted Loci by Methylation.- Ribonuclease Protection.- Quantitative RT-PCR-Based Analysis of Allele-Specific Gene Expression.- Allele-Specific In Situ Hybridization (ASISH).- RNA-FISH to Analyze Allele-Specific Expression.- Flow Cytometry and FISH to Investigate Allele-Specific Replication Timing and Homologous Association of Imprinted Chromosomes.- Southern Analysis Using Methyl-Sensitive Restriction Enzymes.- A PCR-Based Method for Studying DNA Methylation.- Bisulfite-Based Methylation Analysis of Imprinted Genes.- Direct Analysis of Chromosome Methylation.- In Vitro Methylation of Predetermined Regions in Recombinant DNA Constructs.- In Vitro Methylation of Specific Regions in Recombinant DNA Constructs by Excision and Religation.- Detection of Methyl?Sensitive DNA?Binding Proteins with Possible Involvement in the Imprinting Phenomenon.- Probing Chromatin Structure with Nuclease Sensitivity Assays.- Examining Histone Acetlylation at Specific Genomic Regions.- Purification of the MeCP2/Histone Deacetylase Complex from Xenopus laevis.- Reconstitution of Chromatin In Vitro.- Genomic Imprinting in Plants.
Inhaltsverzeichnis
Generation of Monoparental Embryos for Investigation into Genomic Imprinting
Wendy L. Dean, Gavin Kelsey, and Wolf Reik
Deriving and Propagating Mouse Embryonic Stem Cell Lines for Studying Genomic Imprinting
Jeffrey R. Mann
Balanced Translocations for the Analysis of Imprinted Regions of the Mouse Genome
Anne C. Ferguson-Smith, Maxine Tevendale, Pantelis Georgiades, and Valerie Grandjean
Production of YAC Transgenic Mice by Pronuclear Injection
Justin F.-X. Ainscough, Rosalind M. John, and Sheila C. Barton
A Transgenic Approach to Studying Imprinted Genes: Modified BACs and PACs
Rosalind M. John, Justin F.-X. Ainscough, and Sheila C. Barton
Methylation-Sensitive Genome Scanning
Izuho Hatada and Tsunehiro Mukai
Subtraction-Hybridization Method for the Identification of Imprinted Genes
Fumitoshi Ishino, Yoshimi Kuroiwa, Naoki Miyoshi, Shin Kobayashi, Takashi Kohda, and Tomoko Kaneko-Ishino
Identification of Imprinted Loci by Methylation: Use of Methylation- Sensitive Representational Difference Analysis (Me-RDA)
Rachel J. Smith and Gavin Kelsey
Ribonuclease Protection
Joanne L. Thorvaldsen and Marisa S. Bartolomei
Quantitative RT-PCR-Based Analysis of Allele-Specific Gene Expression
Judith Singer-Sam and Chunguang Gao
Allele-Specific In Situ Hybridization (ASISH)
Rolf Ohlsson, Kristian Svensson, Hengmi Cui, Helena Malmikumpu, and Gail Adam
RNA-FISH to Analyze Allele-Specific Expression
Giovanna Braidotti
Flow Cytometry and FISH to Investigate Allele-Specific Replication Timing and Homologous Association of Imprinted Chromosomes
Janine LaSalle and Marc Lalande
Southern Analysis Using Methyl-Sensitive Restriction Enzymes
Tom Moore
A PCR-Based Method for Studying DNA Methylation
Mira Ariel
Bisulfite-Based Methylation Analysis of Imprinted Genes
Sabine Engemann, Osman El-Maarri, Petra Hajkova, Joachim Oswald, and Joern Walter
Direct Analysis of Chromosome Methylation
Déborah Bourc'his and Evani Viegas-Péquignot
In Vitro Methylation of Predetermined Regions in Recombinant DNA Constructs
Ilse Van den Wyngaert, Roger L. P. Adams, and Stefan U. Kass
In Vitro Methylation of Specific Regions in Recombinant DNA Constructs by Excision and Religation
Ghislaine Dell, Marika Charalambous, and Andrew Ward
Detection of Methyl-Sensitive DNA-Binding Proteins with Possible Involvement in the Imprinting Phenomenon
Kerstin Otte and Björn Rozell
Probing Chromatin Structure with Nuclease Sensitivity Assays
Richard I. Gregory, Sanjeev Khosla, and Robert Feil
Examining Histone Acetylation at Specific Genomic Regions
Ji-Fan Hu and Andrew R. Hoffman
Purification of the MeCP2/Histone Deacetylase Complex from Xenopus laevis
Peter L. Jones, Paul A. Wade, and Alan P. Wolffe
Reconstitution of Chromatin In Vitro
Kiyoe Ura and Yasufumi Kaneda
Genomic Imprinting in Plants
Rinke Vinkenoog, Melissa Spielman, Sally Adams, Hugh G. Dickinson, and Rod J. Scott
Klappentext
Genomic imprinting is the process by which gene activity is regulated according to parent of origin. Usually, this means that either the maternally inherited or the paternally inherited allele of a gene is expressed while the opposite allele is repressed. The phenomenon is largely restricted to mammals and flowering plants and was first recognized at the level of whole genomes. Nuclear transplantation experiments carried out in mice in the late 1970s established the non-equivalence of the maternal and paternal genomes in mammals, and a similar conclusion was drawn from studies of interploidy crosses of flowering plants that extend back to at least the 1930s. Further mouse genetic studies, involving animals carrying balanced translocations (reviewed in Chapter 3), indicated that imprinted genes were likely to be widely scattered and would form a minority within the mammalian genome. The first imprinted genes were identified in the early 1990s; over forty are now known in mammals and the list continues steadily to expand.
Includes supplementary material: sn.pub/extras
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