| Introduction | 1 | | |
| What is genetic engineering? | 2 | | |
| Genes and Genome | 3 | | |
| The flow of genetic information | 4 | | |
| Molecular cloning | 5 | | |
| Basic techniques | 6 | | |
| Basic Techniques of Gene Manipulation-an overview | 7 | | |
| Isolation of genomic DNA | 8 | | |
| Agarose gel electrophoresis | 9 | | |
| Nucleic acid blotting | 10 | | |
| Autoradiography | 11 | | |
| Southern blotting | 12 | | |
| Northern blotting | 13 | | |
| Western blotting | 14 | | |
| Transformation of E. coli , Electroporation | 15 | | |
| Transformation with other organisms | 16 | | |
| Polymerase Chain Reaction (PCR) | 17 | | |
| Cutting and Joining DNA molecules | 18 | | |
| Host-controlled restriction modification | 19 | | |
| Types of restriction and modification (R-M) system | 20 | | |
| Nomenclature of restriction endonucleases | 21 | | |
| Target sites of restriction endonucleases | 22 | | |
| Number and size of restriction fragments | 23 | | |
| Summary of restriction endonucleases | 24 | | |
| DNA modifying enzymes | 25 | | |
| Methods of joining DNA fragments | 26 | | |
| DNA ligase to create covalent recombinant DNA | 27 | | |
| Alkaline Phosphatase | 28 | | |
| Blunt end ligation via linker molecules | 29 | | |
| Adaptors | 30 | | |
| Homopolymer tailing | 31 | | |
| Cloning of cDNA by Homopolymer tailing | 32 | | |
| Plasmids | 33 | | |
| Interconversion of plasmid DNA | 34 | | |
| Effect of ethidium bromide on supercoiling of DNA | 35 | | |
| Phenotypic traits exhibited by plasmids | 36 | | |
| Properties of Conjugative and non-conjugative plasmids | 37 | | |
| Host range of plasmids | 38 | | |
| Partitioning and segregative stability of plasmids | 39 | | |
| Incompatibility of plasmids | 40 | | |
| Purification of plasmid DNA | 41 | | |
| Desirable properties of plasmid cloning vehicles | 42 | | |
| Natural plasmids as cloning vehicles | 43 | | |
| Use of pSC101 for cloning | 44 | | |
| pBR322, a purpose-built cloning vehicle | 45 | | |
| Cloning with pBR322 | 46 | | |
| Improved vectors derived from pBR322 | 47 | | |
| Runaway plasmid vectors | 48 | | |
| Bacteriophage lambda as cloning vector | 49 | | |
| Replication of phage-lambda DNA in lytic and lysogenic cycles | 50 | | |
| Modified lambda phages | 51 | | |
| Steps in cloning with lambda | 52 | | |
| Packaging phage-lambda DNA in vitro | 53 | | |
| Vectors for DNA sequencing: bacteriophage M13 | 54 | | |
| Cosmid vectors | 55 | | |
| Modified schemes for cloning in Cosmid vectors | 56 | | |
| Phasmid vectors | 57 | | |
| Bacterial artificial chromosomes (BACs) | 58 | | |
| Yeast artificial chromosomes (YACs) | 59 | | |
| Shuttle and Expression vectors | 60 | | |
| Comparison of different cloning vectors-summary | 61 | | |
| Overview of cloning strategies | 62 | | |
| Genomic DNA libraries | 63 | | |
| LambdaEMBL vectors for genomic library construction | 64 | | |
| Genomic libraries in high-capacity vectors | 65 | | |
| PCR as an alternative to genomic DNA cloning | 66 | | |
| Properties of cDNA | 67 | | |
| cDNA libraries | 68 | | |
| Preparation of cDNA for cloning | 69 | | |
| Improved methods for cDNA cloning | 70 | | |
| PCR as an alternative for cDNA cloning | 71 | | |
| Screening strategies | 72 | | |
| Screening by hybridization | 73 | | |
| Benton and Davis’ plaque lift procedure | 74 | | |
| Probe design | 75 | | |
| Chromosome walking | 76 | | |
| Chromosome jumping | 77 | | |
| Screening by PCR | 78 | | |
| Expression cloning | 79 | | |
| Immunochemical screening | 80 | | |
| Immunochemical screening of lambdagt11 | 81 | | |
| South-western and north-western blotting | 82 | | |
| Screening by functional complementation | 83 | | |
| Requirement for expression in E. coli | 84 | | |
| Secretion of proteins | 85 | | |
| Protein trafficking | 86 | | |
| Stability of foreign proteins in E. coli | 87 | | |
| Constructing the optimal promoter | 88 | | |
| Optimizing translation initiation | 89 | | |
| Stability of mRNA and codon choice | 90 | | |
| The effect of plasmid copy number | 91 | | |
| Plasmid stability | 92 | | |
| Structural instability | 93 | | |
| Host cell physiology can affect the level of expression | 94 | | |
| DNA sequencing: Benefits and Applications | 95 | | |
| Maxam-Gilbert method | 96 | | |
| Chain termination or dideoxy procedure | 97 | | |
| Modifications of chain terminator sequencing | 98 | | |
| Automated DNA sequencing | 99 | | |
| Sequencing accuracy | 100 | | |
| DNA sequence databases | 101 | | |
| Mutagenesis | 102 | | |
| Cassette mutagenesis | 103 | | |
| Primer extension: the single primer method | 104 | | |
| PCR methods for site-directed mutagenesis | 105 | | |
| Basic PCR reaction | 106 | | |
| PCR principles and procedure | 107 | | |
| DNA polymerases | 108 | | |
| Primers | 109 | | |
| Degenerate primers | 110 | | |
| Types of PCR | 111 | | |
| Competitor RT-PCR | 112 | | |
| Real-time quantitative PCR (qPCR) | 113 | | |
| Nested PCR | 114 | | |
| Inverse PCR | 115 | | |
| Multiplex PCR | 116 | | |
| RAPD | 117 | | |
| RFLP | 118 | | |
| AFLP-PCR | 119 | | |
| Applications of PCR | 120 | | |
| PCR-Medicine | 121 | | |
| PCR-Forensic sciences | 122 | | |
| PCR-Agricultural sciences and environment | 123 | | |
| PCR-Molecular paleontology | 124 | | |
| Genome mapping | 125 | | |
| Markers for genome mapping | 126 | | |
| Genetic linkage mapping | 127 | | |
| Physical mapping | 128 | | |
| Physical versus linkage maps | 129 | | |
| The use of RFLPs in physical maps | 130 | | |
| STS in physical maps | 131 | | |
| SNPs as physical markers | 132 | | |
| Polymorphic DNA detection in the absence of sequence information | 133 | | |
| AFLPs detection in the absence of sequence information | 134 | | |
| Fluorescence in situ hybridization (FISH) | 135 | | |
| Radiation hybrid (RH) mapping | 136 | | |
| Happy mapping | 137 | | |
| Map integration | 138 | | |
| Sequencing Genome | 139 | | |
| High-throughput sequencing | 140 | | |
| Shotgun sequencing | 141 | | |
| Clone-by-clone sequencing | 142 | | |
| Orthologs and paralogs | 143 | | |
| Comparative genomics of bacteria | 144 | | |
| Comparative genomics of organelles | 145 | | |
| Comparative genomics of eukaryotes | 146 | | |
| DNA Microarray | 147 | | |
| Spotted DNA Arrays | 148 | | |
| Oligonucleotide Chips | 149 | | |
| Applications of Microarray-Microbial gene expression analysis | 150 | | |
| Applications of Microarray-Profiling in human disease | 151 | | |
| Phage Display | 152 | | |
| Screening phage display libraries | 153 | | |
| Applications of phage display | 154 | | |
| Knock Outs & Knock Inns | 155 | | |
| siRNA Technology | 156 | | |
| Applications of siRNA | 157 | | |
| Proteomics | 158 | | |
| Two-dimensional (2-D) electrophoresis | 159 | | |
| Mass Spectrometry | 160 | | |
| Protein Microarrays | 161 | | |
| Cloning in gram –ve other than E. coli | 162 | | |
| Cloning in gram +ve bacteria B. subtilis | 163 | | |
| Cloning in Streptomyces | 164 | | |
| Cloning in Archaea | 165 | | |
| Cloning in S. cerevisiae | 166 | | |
| Transformation of fungi with exogenous DNA | 167 | | |
| Vectors for use in S. cerevisiae | 168 | | |
| Properties of different yeast vectors | 169 | | |
| Promoter system for yeast | 170 | | |
| Multipurpose vectors for use in yeast | 171 | | |
| Cloning of large DNA fragments | 172 | | |
| Deficiencies and advantages of YACs | 173 | | |
| Plant transformation | 174 | | |
| Callus cultrue | 175 | | |
| Cell suspension culture | 176 | | |
| Protoplast culture | 177 | | |
| Regenration of fertile plants | 178 | | |
| Major strategies for transformation | 179 | | |
| Topic_180_ Agrobacterium mediated transformation | 180 | | |
| Topic_181_T-DNA tansfer | 181 | | |
| Topic_182_Function of T-DNA gene transfer | 182 | | |
| Topic_183_Disarmed Ti vectors | 183 | | |
| Topic_184_Two plasmid strategy | 184 | | |
| Topic_185_Leaf disc transformation | 185 | | |
| Topic_186_Agrobacterium and monocots | 186 | | |
| Topic_187_Rhizogenes and Ri plasmids | 187 | | |
| Topic_188_Protoplast transformation | 188 | | |
| Topic_189_Particle bombardment | 189 | | |
| Topic_190_Chloroplast transformation | 190 | | |
| Topic_191_In planta transformation | 191 | | |
| Topic_192_Plant viruses as vectors | 192 | | |
| Topic_193_Genetic modification in animals | 193 | | |
| Topic_194_Strategies to transform animal cells | 194 | | |
| Topic_195_Chemical transfection techniques | 195 | | |
| Topic_196_Transfection with polyplexes | 196 | | |
| Topic_197_Transfection with liposomes and lipoplexes | 197 | | |
| Topic_198_Physical transfection techniques | 198 | | |
| Topic_199_Microinjections | 199 | | |
| Topic_200_Selectable markers for animal cell gene transfer | 200 | | |
| Topic_201_Bacterial vectors for animal transfection | 201 | | |
| Topic_202_Vrial vectors for transfection | 202 | | |
| Topic_203_Transgenic mice production | 203 | | |
| Topic_204_Pronuclear microinjection | 204 | | |
| Topic_205_Recombinant retroviruses | 205 | | |
| Topic_206_Transfection ES cells | 206 | | |
| Topic_207_Application of transgenic mice | 207 | | |
| Topic_208_Gene transfer to Xenopus | 208 | | |
| Topic_209_Gene transfer to Fish | 209 | | |
| Topic_210_Ways to exploit recombinant DNA | 210 | | |
| Topic_211_Production of recombinant proteins | 211 | | |
| Topic_212_Biopharmaceuticals approved | 212 | | |
| Topic_213_Transgenic animals and plants | 213 | | |
| Topic_214_Recombinant proteins in plants | 214 | | |
| Topic_215_Metabolic engineering in bacteria | 215 | | |
| Topic_216_Metabolic engineering in plants | 216 | | |
| Topic_217_Herbicide resistance in plants | 217 | | |
| Topic_218_Recombinant virus production | 218 | | |
| Topic_219_Fungal resistance | 219 | | |
| Topic_220_Resistance to bacteria | 220 | | |
| Topic_221_BT an insect resistance gene source | 221 | | |
| Topic_222_Drought resistance transgenic crops | 222 | | |
| Topic_223_Transgenic crops for poor quality soil | 223 | | |
| Topic_224_Biotechnology role in food | 224 | | |
| Topic_225_Transgenic model animals | 225 | | |
| Topic_226_Gene medicine | 226 | | |
| Topic_227_DNA vaccines | 227 | | |
| Topic_228_Gene augmentation therapy | 228 | | |
| Topic_229_Gene therapy in cancer | 229 | | |
| Topic_230_Ethics in gene manipulation | 230 | | |