GENETIC ENGINEERING syllabus, JNTU (2007-2008)

Unit I: Gene Regulation and Expression in Prokaryotes

Lactose, Arabinose and Tryptophan operons, Repressors and activator, Sigma switch in Bacillus subtilis.

Unit II: Gene Regulation in Eukaryotic system

Gene regulation in Eukaryotic system, Repetitive DNA, Gene rearrangement, Promoters, enhancer elements, gene amplification.

Unit III: Plasmids, Transposons / Vectors for Gene Transfers

Plasmids: Definition, types, Identification, classification and purifications and transfer of Plasmids. Host restriction in transfer. Transposable elements: Definition, detection of transposition in bacteria, types of

bacterial transposons, mechanisms of transposition and excision, applications of

transposons. Retrotransposons.

UNIT IV: DNA Technology

Purification of genomic DNA from living cells, Manipulation of purified DNA; construction of prototype vector (pBR 322), different types of cloning vectors (plasmid – pUC 19, ? phage, cosmid, M13). Enzymes involved in genetic engineering; cloning

strategies, Introduction of DNA into living cells. Methods of Gene transfer, Restriction mapping.

UNIT – V Expression and Detection of clones

Detection of clones and its expression: Expression of cloned genes in yeast & E. coli. Blot analysis - Southern, Northern & Western blot; dot and slot blot. Immunological techniques. DNA methylation, DNA hybridization. Genomic and cDNA library construction and application. DNA sequencing.

Unit VI: PCR and its application

Principles, designing of primers, PCR methodology, RT - PCR, multiplex PCR, identification of PCR product, application of PCR technology.

Unit VII: Molecular markers

Molecular markers: RFLP, RAPD, AFLP, 16s r-RNA typing, gene chip and micro array; applications in disease profile

Unit VIII: Applications of r-DNA Technology

Gene cloning in medicine (Insulin, Blood clotting factor VIII) High level expression of proteins in different host systems (E. coli, yeast, Insect, mammalian cells) Limitation and advantages and novel technologies- for generation of transgenic animals. Introduction to Gene therapy (Ex vivo & In vivo), case study of ADA as an example. Advantages and limitations of Gene therapy.

TEXT BOOKS:

1. Old RW, Primrose SB, principles of Gene manipulation, An introduction to Genetic engineering, Blackwell Scientific Publications, 1993

2. T.A. Brown, Gene Cloning.

REFERENCES:

1. Ansubel FM., Brent A, Kingston AE, Moore DO, Current protocols in Molecular Biology, Greene Publishing Associates, NY, 1988.

2. Berger SL, Kimmer AR, Methods in Enzymology, Vol 152, Academic Press, 1987.

3. Molecular Cell Biology – Gerald Carp.