Methods in molecular biology

著者

• Walker, John M.

書誌事項

Methods in molecular biology

edited by John M. Walker

（Biological methods）

Humana Press, c1984-c1988

• v. 1 : Proteins
• v. 1 : Proteins : pbk
• v. 2 : Nucleic acids
• v. 2 : Nucleic acids : pbk
• v. 3 : New protein techniques
• v. 4 : New nucleic acid techniques

内容説明・目次

巻冊次

v. 1 : Proteins ISBN 9780896030626

内容説明

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "'genetic en- neering". While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

目次

The Lowry Method for Protein Quantitation.- Determination of Protein Molecular Weights by Gel Permeation High Pressure Liquid Chromatography.- Immunoaffinity Purification of Protein Antigens.- Electrophoretic and Chromatographic Separation of Peptides on Paper.- Peptide Mapping by Reverse-Phase High Pressure Liquid Chromatography.- SDS Polyacrylamide Gel Electrophoresis of Proteins.- Gradient SDS Polyacrylamide Gel Electrophoresis.- Acetic Acid-Urea Polyacrylamide Gel Electrophoresis of Proteins.- The In Vivo Isotopic Labeling of Proteins for Polyacrylamide Gel Electrophoresis.- Two-Dimensional Polyacrylamide Gel Electrophoresis of Proteins.- Starch Gel Electrophoresis of Proteins.- Isoelectric Focusing in Ultrathin Polyacrylamide Gels.- High-Sensitivity Silver Staining of Proteins Following Polyacrylamide Gel Electrophoresis.- Quantification of Proteins on Polyacrylamide Gels (Nonradioactive).- Computer Analysis of Gel Scans.- Drying Gels.- Fluorography of Polyacrylamide Gels Containing Tritium.- Recovery of Proteins from Dried Polyacrylamide Gels after Fluorography.- The Electrophoretic Elution of Proteins from Polyacrylamide Gels.- Transfer Techniques in Protein Blotting.- Peptide Mapping by Thin-Layer Chromatography and High Voltage Electrophoresis.- In Situ Peptide Mapping of Proteins Following Polyacrylamide Gel Electrophoresis.- The Dansyl Method for Identifying N-Terminal Amino Acids.- The Dansyl-Edman Method for Peptide Sequencing.- Microsequencing of Peptides and Proteins with 4-N,N-Dimethylaminoazobenzene-4?-isothiocyanate.- Manual Edman Degradation of Proteins and Peptides.- Carboxyl-Terminal Sequence Determination of Proteins and Peptides with Carboxypeptidase Y.- Immunization and Fusion Protocols for Hybridoma Production.- Immunofluorescence and Immunoperoxidase Screening of Hybridomas.- Solid-Phase Screening of Monoclonal Antibodies.- Subclass Analysis and Purification of Monoclonal Antibodies.- The Production of Antisera.- Immunodiffusion in Gels.- Crossed Immunoelectrophoresis.- Rocket Immunoelectrophoresis.- Radioiodination of Proteins.- Radioimmunoassay.- Enzyme-Linked Immunosorbant Assay (ELISA).

巻冊次

v. 2 : Nucleic acids ISBN 9780896030640

内容説明

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "genetic en- neering". While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

目次

The Burton Assay for DNA.- DABA Fluorescence Assay for Submicrogram Amounts of DNA.- Preparation of "RNase-Free" DNase by Alkylation.- The Isolation of Satellite DNA by Density Gradient Centrifugation.- The Isolation of High Molecular Weight Eukaryotic DNA.- Preparation of Lyophilized Cells to Preserve Enzyme Activities and High Molecular Weight Nucleic Acids.- Agarose Gel Electrophoresis of DNA.- Autoradiography of Gels Containing 32P.- Detection of Specific DNA Sequences-The Southern Transfer.- The Extraction and Isolation of DNA from Gels.- One-Dimensional Electrophoresis of Nucleic Acids in Agarose Using Denaturation with Formaldehyde and Identification of 3H-Labeled RNA by Fluorography.- Gel Electrophoresis of RNA in Agarose and Polyacrylamide Under Nondenaturing Conditions.- The Extraction of Total RNA by the Detergent and Phenol Method.- RNA Extraction by the Proteinase K Method.- RNA Extraction by the Guanidine Thiocyanate Procedure.- The Purification of Poly(A)-Containing RNA by Affinity Chromatography.- Messenger RNA Fractionation on Neutral Sucrose Gradients.- The Estimation of mRNA Content by Poly(U) Hybridization.- DNA Directed in Vitro Protein Synthesis with Escherichia coli S-30 Extracts.- In Vitro Translation of Messenger RNA in a Wheat Germ Extract Cell-Free System.- In Vitro Translation of Messenger RNA in a Rabbit Reticulocyte Lysate Cell-Free System.- Immunoprecipitation of in Vitro Translation Products with Protein A Bound to Sepharose.- In Vitro Continuation of RNA Synthesis Initiated in Vivo.- Synthesis of Double-Stranded Complementary DNA from Poly(A)+mRNA.- Plasmid DNA Isolation by the Cleared Lysate Method.- Plasmid DNA Isolation (Sheared Lysate Method).- Small-Scale Plasmid DNA Preparation.- Preparation of Chromosomal DNA from E. coli.- Preparation and Assay of Phage Lambda.- Preparation of Phage Lambda DNA.- The Use of Restriction Endonucleases.- Ligation of DNA with T4 DNA Ligase.- The Use of Alkaline Phosphatase to Prevent Vector Regeneration.- Bacterial Transformation.- Bacterial Transformation (Kushner Method).- In Vitro Packaging of DNA.- Yeast Transformation.- Radiolabeling of DNA by Nick Translation.- Radiolabeling of DNA Using Polynucleotide Kinase.- Radiolabeling of DNA with 3? Terminal Transferase.- Radiolabeling of DNA with the Klenow Fragment of DNA Polymerase.- Identification of Recombinant Plasmids by In Situ Colony Hybridization.- Identification of Recombinant Phages by Plaque Hybridization.- Plasmid Screening Using Single Colony Lysates.- Immunological Detection of Gene Expression in Recombinant Clones.- The Isolation of Minicells.- The Isolation of Maxicells.- The Identification of Gene Products in Minicells and Maxicells.- Molecular Cloning in Bacteriophage Lambda and in Cosmids.- DNA Transformation of Mammalian Cells.- Chemical Cleavage (Maxam and Gilbert) Method for DNA Sequence Determination.- Gel Electrophoretic Analysis of DNA Sequencing Products.- DNA Sequence Determination Using Dideoxy Analogs.

巻冊次

v. 1 : Proteins : pbk ISBN 9780896031067

内容説明

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "'genetic en- neering". While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

目次

The Lowry Method for Protein Quantitation.- Determination of Protein Molecular Weights by Gel Permeation High Pressure Liquid Chromatography.- Immunoaffinity Purification of Protein Antigens.- Electrophoretic and Chromatographic Separation of Peptides on Paper.- Peptide Mapping by Reverse-Phase High Pressure Liquid Chromatography.- SDS Polyacrylamide Gel Electrophoresis of Proteins.- Gradient SDS Polyacrylamide Gel Electrophoresis.- Acetic Acid-Urea Polyacrylamide Gel Electrophoresis of Proteins.- The In Vivo Isotopic Labeling of Proteins for Polyacrylamide Gel Electrophoresis.- Two-Dimensional Polyacrylamide Gel Electrophoresis of Proteins.- Starch Gel Electrophoresis of Proteins.- Isoelectric Focusing in Ultrathin Polyacrylamide Gels.- High-Sensitivity Silver Staining of Proteins Following Polyacrylamide Gel Electrophoresis.- Quantification of Proteins on Polyacrylamide Gels (Nonradioactive).- Computer Analysis of Gel Scans.- Drying Gels.- Fluorography of Polyacrylamide Gels Containing Tritium.- Recovery of Proteins from Dried Polyacrylamide Gels after Fluorography.- The Electrophoretic Elution of Proteins from Polyacrylamide Gels.- Transfer Techniques in Protein Blotting.- Peptide Mapping by Thin-Layer Chromatography and High Voltage Electrophoresis.- In Situ Peptide Mapping of Proteins Following Polyacrylamide Gel Electrophoresis.- The Dansyl Method for Identifying N-Terminal Amino Acids.- The Dansyl-Edman Method for Peptide Sequencing.- Microsequencing of Peptides and Proteins with 4-N,N-Dimethylaminoazobenzene-4?-isothiocyanate.- Manual Edman Degradation of Proteins and Peptides.- Carboxyl-Terminal Sequence Determination of Proteins and Peptides with Carboxypeptidase Y.- Immunization and Fusion Protocols for Hybridoma Production.- Immunofluorescence and Immunoperoxidase Screening of Hybridomas.- Solid-Phase Screening of Monoclonal Antibodies.- Subclass Analysis and Purification of Monoclonal Antibodies.- The Production of Antisera.- Immunodiffusion in Gels.- Crossed Immunoelectrophoresis.- Rocket Immunoelectrophoresis.- Radioiodination of Proteins.- Radioimmunoassay.- Enzyme-Linked Immunosorbant Assay (ELISA).

巻冊次

v. 2 : Nucleic acids : pbk ISBN 9780896031074

内容説明

In recent years there has been a tremendous increase in our understanding of the functioning of the cell at the molecular level. This has been achieved in the main by the invention and development of new methodology, parti- larly in that area generally referred to as "genetic en- neering". While this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same time developed fresh methodology to keep pace with the requirements of present day molecular bi- ogy. Today's molecular biologist can no longer be content with being an expert in one particular area alone. He/she needs to be equally competent in the laboratory at h- dling DNA, RNA, and proteins, moving from one area to another as required by the problem he/she is trying to solve. Although many of the new techniques in molecular biology are relatively easy to master, it is often difficult for a researcher to obtain all the relevant information nec- sary for setting up and successfully applying a new te- nique. Information is of course available in the research l- erature, but this often lacks the depth of description that the new user requires. This requirement for in-depth pr- tical details has become apparent by the considerable - mand for places on our Molecular Biology Workshops held at Hatfield each summer.

目次

The Burton Assay for DNA.- DABA Fluorescence Assay for Submicrogram Amounts of DNA.- Preparation of "RNase-Free" DNase by Alkylation.- The Isolation of Satellite DNA by Density Gradient Centrifugation.- The Isolation of High Molecular Weight Eukaryotic DNA.- Preparation of Lyophilized Cells to Preserve Enzyme Activities and High Molecular Weight Nucleic Acids.- Agarose Gel Electrophoresis of DNA.- Autoradiography of Gels Containing 32P.- Detection of Specific DNA Sequences-The Southern Transfer.- The Extraction and Isolation of DNA from Gels.- One-Dimensional Electrophoresis of Nucleic Acids in Agarose Using Denaturation with Formaldehyde and Identification of 3H-Labeled RNA by Fluorography.- Gel Electrophoresis of RNA in Agarose and Polyacrylamide Under Nondenaturing Conditions.- The Extraction of Total RNA by the Detergent and Phenol Method.- RNA Extraction by the Proteinase K Method.- RNA Extraction by the Guanidine Thiocyanate Procedure.- The Purification of Poly(A)-Containing RNA by Affinity Chromatography.- Messenger RNA Fractionation on Neutral Sucrose Gradients.- The Estimation of mRNA Content by Poly(U) Hybridization.- DNA Directed in Vitro Protein Synthesis with Escherichia coli S-30 Extracts.- In Vitro Translation of Messenger RNA in a Wheat Germ Extract Cell-Free System.- In Vitro Translation of Messenger RNA in a Rabbit Reticulocyte Lysate Cell-Free System.- Immunoprecipitation of in Vitro Translation Products with Protein A Bound to Sepharose.- In Vitro Continuation of RNA Synthesis Initiated in Vivo.- Synthesis of Double-Stranded Complementary DNA from Poly(A)+mRNA.- Plasmid DNA Isolation by the Cleared Lysate Method.- Plasmid DNA Isolation (Sheared Lysate Method).- Small-Scale Plasmid DNA Preparation.- Preparation of Chromosomal DNA from E. coli.- Preparation and Assay of Phage Lambda.- Preparation of Phage Lambda DNA.- The Use of Restriction Endonucleases.- Ligation of DNA with T4 DNA Ligase.- The Use of Alkaline Phosphatase to Prevent Vector Regeneration.- Bacterial Transformation.- Bacterial Transformation (Kushner Method).- In Vitro Packaging of DNA.- Yeast Transformation.- Radiolabeling of DNA by Nick Translation.- Radiolabeling of DNA Using Polynucleotide Kinase.- Radiolabeling of DNA with 3? Terminal Transferase.- Radiolabeling of DNA with the Klenow Fragment of DNA Polymerase.- Identification of Recombinant Plasmids by In Situ Colony Hybridization.- Identification of Recombinant Phages by Plaque Hybridization.- Plasmid Screening Using Single Colony Lysates.- Immunological Detection of Gene Expression in Recombinant Clones.- The Isolation of Minicells.- The Isolation of Maxicells.- The Identification of Gene Products in Minicells and Maxicells.- Molecular Cloning in Bacteriophage Lambda and in Cosmids.- DNA Transformation of Mammalian Cells.- Chemical Cleavage (Maxam and Gilbert) Method for DNA Sequence Determination.- Gel Electrophoretic Analysis of DNA Sequencing Products.- DNA Sequence Determination Using Dideoxy Analogs.

巻冊次

v. 3 : New protein techniques ISBN 9780896031265

内容説明

Inrecent years therehasbeen atremendousincreaseinour understandingofthe functioningofthe cellat the molecular leveL This has been achieved in the main by the invention and developmentof new methodology, particularlyin that area generally referred to as "genetic engineering. " Al- though this revolution has been taking place in the field of nucleic acids research, the protein chemist has at the same timedevelopedfresh methodologytokeeppace with the re- quirements ofpresent-daymolecularbiology. Today's mo- lecularbiologistscannolongerbecontentwithbeingexperts inoneparticulararea alone. Theyneedtobeequallycompe- tentin the laboratory at handling DNA, RNA,and proteins movingfrom one area toanotherasrequiredby theproblem thatisbeing solved. Althoughmanyof thenewtechniques in molecularbiologyare relativelyeasy tomaster, itisoften difficult for a researcher to obtain all the relevant informa- tionnecessaryforsettingupandsuccessfullyapplyinganew technique. Informationisofcourse availablein the research literature, but this often lacks the depth of description that the new user requires. This requirement for in-depth prac- tical details has become apparent by the considerable de- mand for places on our Molecular BiologyWorkshops held at Hatfield each summer. Volume 1of this series describedpracticalprocedures for a range of protein techniques frequently used by research workers in the field of molecular biology. Because of the limitations on length necessarily inherent in producing any v vi Preface book, one obviouslyhad to be selective in the choiceof titles forVolume1. TheproductionofVolume 3,therefore,allows the development of the theme initiated in Volume 1. This volumecontains afurther selection ofdetailed protocols for arangeofanalyticalandpreparativeproteintechniques,and should be seen as a continuation of Volume 1. Companion Volumes2and4provideprotocols fornucleic acid method- ology. Each methodisdescribedby an authorwhohas regularly used the technique in his or her ownlaboratory.

目次

Prevention of Unwanted Proteolysis.- The Bradford Method for Protein Quantitation.- Amino Acid Analysis by Precolumn Derivatization.- Identification of N-Terminal Amino Acids by High-Performance Liquid Chromatography.- Enzymatic Methods for Cleaving Proteins.- Chemical Cleavage of Proteins.- Chemical Modification of Proteins.- The Design, Preparation, and Use of Immunopurification Reagents.- Dye-Ligand Chromatography.- Aminohexyl-Sepharose Affinity Chromatography.- Purification of DNA-Dependent RNA Polymerase from Eubacteria.- Direct Immunoprecipitation of Protein.- Detection of Proteins in Polyacrylamide Gels Using an Ultrasensitive Silver Staining Technique.- Chromatofocusing.- Hybrid Isoelectric Focusing Using Mixed Synthetic-Carrier Ampholyte-Immobilized pH Gradient Gels.- Two-Dimensional Electrophoresis Using Immobilized pH Gradients in the First Dimension.- Two-Dimensional Polyacrylamide Gel Electrophoresis Using Flat-Bed Isoelectric Focusing in the First Dimension.- Preparative Aspects of Immobilized pH Gradients.- Isoelectric Focusing Under Denaturing Conditions.- Computer Analysis of 2-D Electrophoresis Gels.- Two-Dimensional (Crossed) Immunoelectrophoresis.- Peptide Synthesis.- Synthesis of a Series of Analogous Peptides Using T-Bags.- Production of Antisera to Synthetic Peptides.- Production of Antibodies Using Proteins in Gel Bands.- Purification of Immunoglobulins Using Protein A-Sepharose.- Antibody-Enzyme Conjugate Formation.- Vacuum Blotting.- Blotting with Plate Electrodes.- Use of Dried Milk for Immunoblotting.- Immunodetection of Proteins on "Western" Blots Using 125I-Labeled Protein A.- Detection of Protein Blots Using the Avidin-Biotin System.- Detection of Protein Blots Using Enzyme-Linked Second Antibodies or Protein A.- Colloidal Gold for the Detection of Proteins on Blots and Immunoblots.- Enzyme Immobilization by Adsorption.- Enzyme Immobilization by Entrapment.- Enzyme Immobilization by Covalent Bonding.- Cell Immobilization.

巻冊次

v. 4 : New nucleic acid techniques ISBN 9780896031272

目次

Electrophoresis of RNA Denatured with Glyoxal or Formaldehyde.- Northern Blotting.- Hybrid-Release Translation.- Hybrid-Arrested Translation.- In Vitro Translation and Analysis of Early Events in Protein Synthesis Initiation in Nonreticulocyte Mammalian Cells.- Use of Elutips to Purify DNA.- Plasmid Preparation on Sephacryl S1000.- Electrophoresis of DNA in Nondenaturing Polyacrylamide Gels.- Use of 35S Nucleotides for DNA Sequencing.- Direct Dideoxy DNA Sequencing.- Computer Applications to Molecular Biology.- Detection of Sequence-Specific Protein-DNA Interactions by the DNA-Footprinting Technique.- Oligonucleotide Synthesis Using the Manual Phosphotriester Method.- Manual Oligonucleotide Synthesis Using the Phosphoramidite Method.- Purification of Synthetic Oligonucleotides by Preparative Gel Electrophoresis.- Isolation of High Molecular Weight DNA Suitable for the Construction of Genomic Libraries.- Construction of Mammalian Genomic Libraries Using ? Replacement Vectors.- Cosmid Library Construction.- Construction of cDNA Libraries in ?gt10 or ?gt11.- Immunoscreening of ?gt11 Expression Libraries Using an Avidin-Biotin Detection System.- Primed Synthesis and Direct Sequencing in the Isolation of cDNA Clones Using Short Oligonucleotides.- Constructing Expression cDNA Libraries Using Unphosphorylated Adaptors.- Expression Screening of cDNA Libraries in pEX.- Producing Antibodies of Predetermined Specificity from Escherichia coli Hybrid Proteins.- Epitope Mapping Using pEX.- DNA Transfection of Mammalian Cells Using Polybrene.- Expression of Foreign Genes in Mammalian Cells.- Dot-Blot Hybridization Method.- Detection of Unique or Low Copy Number DNA Sequences in Complex Genomes.- Photobiotin-Labeled DNA and RNA Hybridization Probes.- Enzyme-Labeled DNA Probes.- Luminescent Detection of Specific DNA Sequences.- Detection of DNA Sequences Using Biotinylated Probes.- Crosslinking of Single-Stranded DNA to Resins.- Extraction of RNA from Plants.- Isolation of Plant Nuclei.- In Vitro Transcription in Plant Nuclei.- Large-Scale Isolation of Ti Plasmid DNA.- Transformation and Regeneration of Dicotyledonous Plants.- Plant Protoplast Fusion.- Plant Tissue Culture.- Direct Gene Transfer into Plant Protoplasts.- Isolation of Wheat Chloroplasts and Chloroplast DNA.- DNA Labeling by Isolated Wheat Chloroplasts.

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詳細情報

• NII書誌ID(NCID): BA01223853
• ISBN:
  • 0896030628
  • 0896031063
  • 0896030644
  • 0896031071
  • 0896031268
  • 0896031276
• LCCN: 84015696
• 出版国コード: us
• タイトル言語コード: eng
• 本文言語コード: eng
• 出版地: Clifton, N.J.
• ページ数/冊数: v.
• 大きさ: 24 cm
• 分類:
  • LCC : QH506
  • DC19 : 574.8/8/078
• 件名:
  • LCSH : Molecular biology -- Technique -- Collected works
• 親書誌ID:
  • BA00306048