Course

Syllabus

Theory

Impact of Biotechnology on crop improvement and the perspective of society; Various biotechnological techniques available for crop improvement – Plant Tissue Culture, Genetic Engineering, Genome editing, Marker Assisted breeding and Genomic Selection. Biosafety regulations and their application in Agricultural Biotechnology. Somaclonal variation and its use in crop improvement; embryo culture; anther/pollen culture; somatic embryogenesis; artificial seeds; techniques of protoplast culture, regeneration and somatic cell hybridization, achievements and limitations, utility in the improvement of crop plants. Direct and Indirect methods of gene transfer in plants – Agrobacterium-mediated gene transfer in dicots and monocots; Direct DNA delivery methods (microinjection, particle gun method, electroporation); gene targeting; Gene silencing techniques; introduction to siRNA; siRNA technology; Micro RNA; construction of siRNA vectors; principle and application of gene silencing; creation of transgenic plants; debate over GM crops; introduction to methods of genetic manipulation in different model systems. Introduction to genome editing – Various tools of genome editing; CRISPR-Cas9 with specific emphasis on Indian regulations; Cloning genomic targets into CRISPR/Cas9 plasmids; electroporation of Cas9 plasmids into cells; purification of DNA from Cas9 treated cells and evaluation of Cas9 gene editing; in vitro synthesis of single guide RNA (sgRNA); using Cas9/ sgRNA complexes to test for activity on DNA substrates; evaluate Cas9 activity by T7E1 assays and DNA sequence analysis; Applications of CRISPR/cas9 technology in crop plants. Marker Assisted Breeding and Genomic Selection: Introduction to various DNA-based markers and their use in marker-assisted breeding; Foreground Selection, Recombinant Selection and background Selection; Marker-assisted backcross breeding, marker-assisted selection – success stories; Introduction to Genomic Selection.

Practical

Agrobacterium-mediated transformation in Tobacco – preparation of construct, transfer to binary vector, transform Agrobacterium, prepare explant, Inoculation and Co-cultivation, antibiotic based selection of putative transformants, validation using PCR; Genome editing- preparation of CRISPR/CAS construct, direct transfer to plant, analysis of the targets; Planning of a MABB programme – selection of parents, crossing strategies, marker analysis.

Objective

i. To acquaint with biotechnological tools of crop improvement

ii. To know about direct and indirect methods of gene transfer

iii. To introduce about gene editing in plants

iv. To provide knowledge about marker assisted breeding and genomic selection

Course Outcome:

Any student who has undergone the course shall be able to:

Suggested Readings

1. Brown, T A. 2006. Genomes. (3rd Edn.). Garland Science Pub, New York.
2. Gene Cloning and DNA Analysis. 2010. Retrieved from http://biolab.szu.edu.cn/ otherweb/lzc/ genetic%20engineering/courseware/b1.pdf
3. Green M R and Sambrook J. 2012. Molecular Cloning: a Laboratory Manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press.
4. Kumar Pranav and Mina Usha. 2015. Biotechnology: A Problem Approach. Pathfinder Publication.
5. Old R W, Primrose S B and Twyman R M. 2001. Principles of Gene Manipulation and Genomics. (7th Edn.). Oxford: Blackwell Scientific Publications.
6. Ram Hari Har. 2019. Crop Breeding and Biotechnology. Kalyani Publications.
7. Rastogi S C. 2020. Biotechnology: Principles and Applications. Narosa.
8. Sander J D and Joung J K. 2014. CRISPR-Cas systems for Editing, Regulating and Targeting Genomes. Nat Biotechnol. 32:347-355.
9. Singh K H, Kumar Ajay and Parmar Nehanjali. 2019. Agricultural Biotechnology at a Glance. Sharma Publishers & Distributers.
10. Slater. 2008. Plant Biotechnology: The Genetic Manipulation of Plants. Oxford, 400p.