Techniques in Molecular Biology I

Teacher

Dr. Satyabrata Nanda

Category

Core Courses

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Course Name: Techniques in Molecular Biology I

Code (Credit): MMBB0504 (0-3-0)

Course Objectives

- To get a basic overview of molecular biology techniques, good lab practices and recombinant DNA technology.

- To get a hands on training in nucleic acid analysis, protein analysis, chromatography, and bacterial and phage genetics.

CO-PO Mapping

PSO1 : Impart knowledge and understanding of fundamental concepts and techniques of maintaining and enhancing soil fertility, crop production, crop management, crop improvement, biodiversity and sustainability of agriculture worldwide.

PSO2 : Develop the ability to know farming practices and their scope to improve in the rural areas and the environments.

PSO3 : Apply knowledge of basic science through Agricultural microbiology, Plant biochemistry and Biotechnology.

PSO4 : Apply knowledge of Agri-business management and finance to develop profitable agricultural system.

Syllabus

Practical 

Good lab practices, preparation of buffers and reagents. Principle of centrifugation and spectrophotometry. Growth of bacterial culture and preparation of growth curve. Isolation of Genomic DNA from bacteria. Isolation of plasmid DNA from bacteria. Growth of lambda phage and isolation of phage DNA. Isolation of plant DNA (e.g. Rice/Moong/Mango/Merigold). Quantification of DNA by (a) Agarose Gel electrophoresis and (b) Spectrophotometry. PCR using isolated DNA. PAGE Gel electrophoresis. Restriction digestion of plasmid and phage DNA, ligation, Recombinant DNA construction. Transformation of E. coli and selection of transformants. Chromatographic techniques: (a) TLC, (b) Gel Filtration Chromatography, (c) Ion exchange Chromatography, (d) Affinity Chromatography. Dot blot analysis, Southern hybridization, and Northern hybridization. Western blotting and ELISA. Radiation safety and non-radio isotopic procedure.

References 

Sambrook, J., and Russell, R.W (2001) Molecular cloning: A laboratory manual 3rd Edition, Cold spring harbor laboratory press, cold spring harbor, New York.

Wilson, K., and Walker, J., (2018) Principles and Techniques of Biochemistry and Molecular Biology 8th edition, Cambridge University Press.

Ausubel, F. M., Brent, R., Kingston, R. E., Moore, D. D., Seidman, J. G., Smith, J. A.,&Struhl, K., (2002) Short Protocols in Molecular Biology 5th edition, Current Protocols publication.

Session Plan

Session 1

Good lab practices.

Principle        Reference video

Session 2-4

Preparation of buffers and reagents and their sterilization.

Principle        Reference video

Session 5

Principle of centrifugation.

Principle        Reference video

Session 6

Principle of spectrophotometry.

Principle        Reference video 1                Reference video 2

Session 7-10

Growth of bacterial culture and  preparation of bacterial growth curve.

Principle        Reference video

Session 11-12

Isolation of Genomic DNA from bacteria.

Principle        Reference video

Session 13-14

Isolation of plasmid DNA from bacteria.

Principle        Reference video

Session 15-16

Growth of lambda phage.

Principle        Reference video

Session 17-18

Isolation of phage DNA.

Principle        Reference video

Session 19-20

Isolation of plant DNA.

Principle        Reference video

Session 21

Quantification of DNA by Agarose Gel electrophoresis.

Principle        Reference video

Session 22

Quantification of DNA by Spectrophotometry.

Principle        Reference video

Session 23-25

PCR using isolated DNA (RAPD/ISSR/SSR).

Principle        Reference video

Session 26

PAGE Gel electrophoresis.

Principle        Reference video

Session 27-28

Restriction digestion of plasmid.

Principle        Reference video

Session 29-30

Restriction digestion of phage DNA.

Principle        Reference video

Session 31-32

Ligation of DNA.

Principle        Reference video

Session 33-34

Recombinant DNA construction.

Principle        Reference video

Session 35-36

Transformation of E. coli.

Principle        Reference video

Session 37-38

Selection of transformants.

Principle        Reference video

Session 39

Chromatographic techniques - TLC.

Principle        Reference video

Session 40

Chromatographic techniques - Gel Filtration Chromatography.

Principle        Reference video

Session 41

Chromatographic techniques - Ion exchange.

Principle        Reference video

Session 42

Chromatographic techniques - Affinity Chromatography.

Principle        Reference video

Session 43

Dot blot analysis.

Principle        Reference video

Session 44

Southern hybridization.

Principle        Reference video

Session 45

Northern hybridization.

Principle        Reference video

Session 46

Western blotting.

Principle        Reference video

Session 47

ELISA.

Principle        Reference video

Session 48

Radiation safety.

Principle        Reference video

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Dr. Satyabrata Nanda

Associate Professor & Head, Dept. of Biotechnology
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Dr. Satyabrata Nanda has a Ph.D. in Biotechnology with a specialization in Plant Protection. His education includes B-Tech, M-Tech, and Ph.D. in Biotechnology. He’s received the Overseas Postdoctoral Fellowship from the Chinese Academy of Agricultural Sciences for research at China National Rice Research Institute. His research is primarily focused on plant protection by using functional […]