Syllabus
|
Course Code: IN-BTY -801 Course Name: Advanced Recombinant DNA Technology |
||
| MODULE NO / UNIT | COURSE SYLLABUS CONTENTS OF MODULE | NOTES |
|---|---|---|
| 1 | Genomic and cDNA library: Gene library, types and Applications, Making genomic and cDNA libraries in plasmids and phages. PCR product cloning (TA cloning), cDNA synthesis strategies – Linkers – Adapters – Homopolymer tailing; Properties of cDNA, mRNA enrichment Selection of rDNA clones and their expression products: Direct and indirect methods. Drug resistance, gene inactivation, DNA hybridization, Colony and Plaque hybridization; Abundancy probing, Heterologus probing, In-situ hybridization (Southern, Northern and Dot blots and immunological techniques Western blotting), Subtractive hybridization; Protein-Protein interactions - Phage display, Yeast two hybrid system, Yeast three hybrid system. Site Directed Mutagenesis: Oligonucleotide directed mutagenesis, PCR amplified oligonucleotide directed mutagenesis, Random mutagenesis with degenerate oligonucleotide primers / nucleotide analogs. Deletion mutagenesis, Applications |
|
| 2 | Gene expression and Regulation studies: Primer extension, S1 mapping, RNase protection assay, Gel retardation assay, Deletion analysis, Reporter genes, DNA foot printing, Modification interference assays, HRT, HART Manipulation of recombinant gene expression in Prokaryotes: Problems with production of recombinant proteins in E coli, Optimizing expression of foreign genes in E.coli- Strong and regulatory promoters, Codon usage, Fusion proteins, Increasing protein stability and secretion, Translation expression vectors, Protease deficient host strains. Heterologous protein production in Eukaryotes: Saccharomyces cerevisiae and Pistia pastoris expression systems, Baculovirus Insect cell expression systems, Mammalian cell expression system, CRE LOX system and CRISPR/Cas9 Applications of rDNA technology: Diagnostics; Pathogensis; Genetic diversity; Therapeutic proteins-Vaccines. Molecular probes (Production, labelling and uses) |
|
