Analytical techniques in plant science

Title of the Course : Analytical techniques in plant science 

Nature of course : Major/ Core-XV 

Code : BOT-C-15  Total Credits : 04 

Distribution of Marks : 100 [End: 60 (Theory: 45, Pract: 15), In: 40] Total lecture-60 

Course outcomes: 

CO1: Understanding the basic principles of the tools and techniques in plant science. CO2: Ability to apply technologies to solve complex biological questions in Life Sciences.  CO3: Ability to operate sophisticated instruments used in biological science

Learning Outcome: 

1. Describe the instruments used in modern biology 

2. Demonstrate the instruments and techniques used in life sciences. 

Unit 1 : Imaging and related techniques (12 lectures)

Principles of microscopy; Light microscopy; Fluorescence microscopy; Confocal microscopy;  Use of fluorochromes: (a) Flow cytometry (FACS); (b) Applications of fluorescence  microscopy: Chromosome banding, FISH, chromosome painting; Transmission and Scanning  electron microscopy – sample preparation for electron microscopy, cryofixation, negative  staining, shadow casting, freeze fracture, freeze etching. 

Unit 2: Cell fractionation (08 lectures)

Centrifugation: Differential and density gradient centrifugation, sucrose density gradient,  CsCl2gradient, analytical centrifugation, ultracentrifugation, marker enzymes.

Unit 3: Radioisotopes & Spectrophotometry (08 lectures)

Radioisotopes: Use in biological research, auto-radiography, pulse chase experiment.  Spectrophotometry: Principle and its application in biological research.

Unit 4 : Chromatography (08 lectures)

Principle; Paper chromatography; Column chromatography, TLC, GLC, HPLC, Ion-exchange  chromatography; Molecular sieve chromatography; Affinity chromatography.

Unit 5 : Characterization of proteins and nucleic acids (09 lectures)

Mass spectrometry; X-ray diffraction; X-ray crystallography; Characterization of proteins and  nucleic acids; Electrophoresis: AGE, PAGE, SDS-PAGE 

Lab activities

1. Study of Blotting techniques: Southern, Northern and Western, DNA fingerprinting,  DNA sequencing, PCR through photographs. 

2. Demonstration of ELISA. 

3. To separate sugars by thin layer chromatography. 

4. Isolation of chloroplasts by differential centrifugation. 

5. To separate chloroplast pigments by column chromatography. 

6. To estimate protein concentration through Lowry’s methods. 

7. To separate proteins using PAGE. 

8. To separation DNA (marker) using AGE. 

9. Study of different microscopic techniques using photographs/micrographs (freeze fracture,  freeze etching, negative staining, positive staining, fluorescence and FISH).