I= METHYL MODIFICATION IN PROKARYOTIC DNA
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Title of Thesis
METHYL MODIFICATION IN PROKARYOTIC DNA

Author(s)
Anjum Sohail
Institute/University/Department Details
University Of The Punjab/ Centre For Advanced Molecular Biology
Session
1989
Subject
Molecular Biology
Number of Pages
108
Keywords (Extracted from title, table of contents and abstract of thesis)
prokaryotic dna, endonucleases, methylase genes, mismatch repair, protein, strains, plasmids, enzymes, reagents, dcm clones

Abstract
No Abstract

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1182.02 KB
S. No. Chapter Title of the Chapters Page Size (KB)
1 0 Contents
80.1 KB
2 1 General Introduction 1
38.8 KB
3 2 Genetic Restriction And Modification 4
269.03 KB
  2.1 Introduction 4
  2.2 Type I Restriction And Modification Systems 5
  2.3 Type III Restriction And Modification Systems 7
  2.4 Type II Restriction And Modification Systems 8
  2.5 Recognition Specificities Of Restriction Endonucleases 11
  2.6 Type II Specific Methylase 14
  2.7 Cloned Methylase Genes 17
  2.8 Reaction Mechanism 22
4 3 Biological Significance Of DNA Methylation In Escherichia Coli 26
214.13 KB
  3.1 Introduction 26
  3.2 DNA Cytosine Methylation 27
  3.3 DNA Adenine Methylase 29
  3.4 Mismatch Repair In E. Coli 30
  3.5 DAM-Directed Mismatch Repair 30
  3.6 Mismatch Repair During Recombination 32
  3.7 Mismatch Repair In Methylated DNA 33
  3.8 Mismatch Repair And Spontaneous Mutability 34
  3.9 Repair Of Protein – DNA Cross-Links 5-Azacytosine 35
  3.10 Restriction Of DNA By Mar And Mcr Gene Products Of E. Coli 36
  3.11 Regulation Of Gene Expression 37
  3.12 Methylated Bases In Bacteria 42
5 4 General Experimental Techniques
120.46 KB
  4.1 Bacterial Strains And Plasmids 44
  4.2 Enzymes And Reagents 44
  4.3 Preparation Of Chromosomal DNA 45
  4.4 Isolation Of Plasmid DNA 45
  4.5 Cloning Of The Dcm Gene 46
  4.6 Transformation Of Escherichia Coli 47
  4.7 In Vitro Assay Of Dcm Methylase 47
  4.8 In Virto Assay Of The Dcm Methylase 48
  4.9 Quantitation Of Methylase Activity 48
  4.10 Southern Blotting And DNA Hybridization 49
  4.11 Bal- 31 Nuclease Delations 49
  4.12 DNA Sequencing 50
  4.13 35S Labeling Of Protein 51
  4.14 Purification Of Methylase 52
  4.15 Computer Analysis 53
6 5 Results 54
293.02 KB
  5.1 Cloning Of The Dcm Gene In Pbr322 54
  5.2 DCM Clones Modify Ecorii Recogition Sites 54
  5.3 Pdcm1 Can Complement R+M-Mutants Of Ecorii 57
  5.4 Pdcm1 Produces A Methyltransferase 58
  5.5 Localization Of The Methylase Coding Region 60
  5.6 Homology Between Pdcm1, DCM3 And C. Coli Chromosome 62
  5.7 Nucleotide Sequenc And Analysis Of Dcm Gene Locus 62
  5.8 Sequence Analysis Of The Vsr Locus 69
  5.9 DNA Homology Between The Dcm Methylase And The Ecorii Restriction Modification System 71
  5.10 Cloning Of The Dcm Methylase Under The Control Of The T7 RNA Polymerase Promoter 73
  5.11 Expression Of The Dcm Methylase 73
  5.12 Purification Of The Dcm Methylase 73
  5.13 Cloned Dcm Gene Can Complement The Dcm-6 Mutation For VSP Repair 76
  5.14 Homology Between The Dcm Methylase And Other Cytosine Methylases 80
7 6 Discussion 83
47.7 KB
8 7 References 87
274.33 KB