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Colorimetry: Introduction • Principle of Colorimetric and
Photometric Estimations • Beer-Lambert’s Law • Colorimetry •
Methods • Absorption of Radiant Energy • Transmittance •
Photoemissive Cells • Photovoltaic Cells • Double Cell Filter
Photometer
Spectrophotometry: Principle • Construction of a Spectrophotometer
• Source of Radiant Energy • Incandescent Lamp • Hydrogen Discharge
Tube • Mercury Discharge Lamp • Globars and Nernst Glowers •
Monochromators • Prisms • Grating • Slits • Cuvette or Cells •
Construction of a Spectrophotometer • Relationship between
Absorption in Ultraviolet and Visible Range and Molecular Structure
• Determination of Absorption Spectrum and Absorption Maximum
Carbohydrates: Qualitative Tests for Sugars/Polysaccharides •
Molisch Test (-Naphthol Reaction) • Moore’s Test • Fehling’s Test •
Benedict’s Test • Benedict’s Reagent • Barfoed’s Test • Barfoed’s
Reagent • Nylander’s Test • Nylander’s Reagent • Picric Acid Test •
Seliwanoff’s Test (Resorcinol Test) • Seliwanoff’s Reagent • Cole’s
Test • Methylamine Hydrochloride Test (FEARON’S TEST) • Mucic Acid
Test • Alcoholic Fermentation • Iodoform Test • Iodine Test • For
Starch • Estimation of Reducing Sugars Using • Nelson–Somogyi
Method • Estimation of Amount of Blood Sugar Using Nelson–Somogyi
Method • Estimation of Blood Glucose by Folin-Wu Method •
Estimation of Reducing Sugars by Dinitrosalicylic (DNS) Acid Method
• Determination of Keto Sugars Using Roe Nicholas and Papadopoulos
Method • Estimation of Total Carbohydrates Using Rex Montgomrey
(phenol–HSO) Method • Estimation of Total Carbohydrates Using
Anthrone Method • Estimation of Glucose Using Glucose Oxidase,
Peroxidase and O-dianisidine (Trinder Test) • Isolation of Starch
from Potatoes • Hydrolysis of Starch by Amylase (Saliva) •
Isolation of Glycogen from Liver
Amino Acids and Proteins: Introduction • Qualitative Tests •
General Composition Test for Proteins • Test for Organic Nitrogen •
Test for Sulfur • Millon’s Test • Millon-Nasse Test • Xanthoproteic
Test • Hopkins–Cole Test • Biuret Test • Ninhydrin
(Triketohydrindene Hydrate) Reaction • Sakaguchi Test • Isolation
of Crystallized Albumin from Egg • Isolation of Glutenin and
Gliadin from Wheat Flour • Determination of the Isoelectric Point
of Casein • Isolation of Casein from Milk • Quantitative Tests for
Amino Acids/Proteins • Estimation of Protein Using
SpectrophotometricMethod • Estimation of Protein Using
Turbidimetric Method • Estimation of Protein Using Bicinchoninic
Acid (BCA) • Advantages • Estimation of Protein Using Biuret Method
• Estimation of Protein Using Folin-Ciocalteau (Lowry) Method •
Estimation of Tyrosine and Protein in a given Mixture Using
Folin-Ciocalteau Method • Advantages and Disadvantages • Estimation
of Protein Using Bradford Method • Estimation of Amino Acids Using
Ninhydrin Reaction • Estimation of Amino Acid Using Sorensen Formol
Titration • Estimation of Purity of an Amino Acid Sample Using
Sorensen Formal Titration • Estimation of Amino Acid Nitrogen in
Urine Sample Using Sorensen Formal Titration • Determination of pK
of an Amino Acid
Lipids: Introduction • Qualitative Tests • Emulsification • Salting
Out of Soap • Acrolein Test for Glycerol • Salkowski Test for
Cholesterol • Determination of Cholesterol Content by
Liebermann-Burchard Method • Baudin Test for Checking the Purity of
Clarified Butter (Ghee) • Determination of Saponification Number
(Koettes-Dorfers Number) of Fat • Determination of Iodine Number of
Oil/Fat • Determination of Acid Value of Fat • Isolation of
Cholesterol from Egg Yolk
Nucleic Acids: Introduction • Isolation of Chromosomal DNA from
Bacteria • Isolation of DNA from Blood • Isolation of DNA from
Plant Leaf Tissue • Estimation of DNA Concentration Using Diphe
Nylamine Method • Isolation of RNA from Yeast • Isolation of Total
Cellular RNA from Eukaryotes (Plant Tissue) • Estimation of RNA
Concentration Using Orcinol Method • Estimation of Nucleic Acid
(DNA) Using Spectrophotometric Method • Separation of DNA and RNAs
• Separation of RNA, Single-stranded DNA and Double-stranded DNA
using Cesium Chloride Density Gradient Centrifugation • Separation
of Different Types of RNA Using Sucrose Density Gradient
Centrifugation • Separation of Different Size Fragments of DNA or
RNA through Electrophoresis and their Separation from Gel •
Fractionation of mRNA from Total Cellular RNA • Isolation of
Plasmid DNA (Miniprep) • Large-Scale Preparation of Plasmid DNA •
Digestion of Plasmid DNA with Restriction Endonucleases
Vitamins: Introduction • Estimation of Vitamin C (Ascorbic Acid)
Using ,-dichlorophenolindophenol (DCPIP) Dye Titration • Estimation
of Ascorbic Acid in a Tissue • Determination of Ascorbic Acid
Saturation in Body • Estimation of Thiamine by Thiochrome
Technique
Nitrogen: Introduction • Estimation of Inorganic Nitrogen Using •
Nessler’s Method • Estimation of Nitrogen in the Presence of Carbon
Compounds by Micro-Kjeldahl Method • (A) Digestion • (B)
Distillation
Phosphorus: Introduction • Estimation of Inorganic Phosphorus Using
Fiske and Subbarow Method • Estimation of Inorganic Phosphorus
Using Sumner Method • Estimation of Inorganic Phosphorus Using
King’s Method • Estimation of Inorganic Phosphorus Using Lowry and
Lopez Method • Estimation of Inorganic Phosphorus Using Berenblum
and Chain Method Modified by Martin and Doty • Estimation of
Organic Phosphorus
Enzymes: Introduction • Determination of the Activity of Acid
Phosphatase from Cuscuta Reflexa • Determination of Activity of
Acid Phosphatase from Germinating Lentil • Effect of Time on the
Activity of Acid Phosphatase • Effect of Enzyme Concentration on
the Activity of Acid Phosphatase • Determination of Optimum pH for
Acid Phosphatase from Cuscuta Reflexa • Determination of Km and
Vmax (Maximal Velocity) Values of Acid Phosphatase for Its
Substrate, Glycerophosphate • Determination of Optimum Temperature
for Acid Phosphatase from Cuscuta Reflexa • Determination of
Inhibition Constant (Ki) of Acid Phosphatase for an Inhibitor
(Fluoride) Using Dixon Plot • Fractionation of Acid Phosphatase
Using Ammonium Sulfate • Determination of Activity of Starch •
Phosphorylase from Banana Leaves/Indian Millet Seeds Using
Colorimetry/Spectrophotometry • Determination of the Activity of
Pectin Methylesterase from Vigna Sinensis Seeds Using an UV-VIS
Spectrophotometer • Study the Synthesis of Amylase During
Germination of Bengal Gram (Chickpea) Seeds • Enzyme Assay •
Immobilization of Enzymes • Methods for Immobilization of Enzymes •
I Carrier-binding Method • IA Physical Adsorption Method •
Immobilization of Enzyme Starch Phosphorylase Isolated from Sorghum
Leaves on Alumina by Physical Adsorption • Immobilization of Enzyme
Starch Phosphorylase Isolated from Cabbage Leaves on Egg Shell by
Physical Adsorption • Immobilization of Enzyme Pectin
Methylesterase Isolated from Vigna Sinensis Seeds on Egg Shell by
Physical Adsorption • IB Ionic Binding Method • Immobilization of
Enzyme Starch Phosphorylase Isolated from Bengal Gram Seeds on
DEAE-cellulose by Ionic Binding • IC Covalent Binding Method • II
Cross-linking Method • Immobilization of Enzyme Starch
Phosphorylase Isolated from Sorghum Leaves on Egg Shell Using
Glutaraldehyde as Cross-linking Reagent • Immobilization of Enzyme
Endogalacturonase Isolated from Vigna Sinensis Seeds on Sand Using
Glutaraldehyde as Cross-linking Reagent • Calculation of Enzyme
Activity • III Entrapping Method • Immobilization of Enzyme Pectin
Methylesterase Isolated from Vigna Sinensis in Gelatin • IIIA
Lattice Type • Immobilization of Enzyme Starch Phosphorylase
Isolated from Sorghum Leaves in Polyacrylamide • IIIB
Microencapsulated Type • Enzyme Membranes • Enzyme Tubes •
Advantages of Immobilized Enzymes • Applications of Immobilized
Enzymes
Milk Analysis: Introduction • Determination of Specific Gravity of
Milk • Determination of pH of Milk • Determination of pH of Milk
Using pH Paper • Determination of pH of Milk Using pH Meter •
Isolation of Proteins from Milk • Detection of Electrolytes
(Calcium and Phosphate) in Milk • Test for Calcium Ions • Test for
Phosphate • Detection of Common Adulterants in Milk • Detection of
Sucrose in Milk • Detection of Starch in Milk by Iodine Test •
Quality Test (Presence of Microbial Cells) of Milk by Methylene
Blue Dye Reduction Test • Determination of Total Solids in Milk •
Determination of Fat Content of Milk
Pigments: Estimation of Chlorophyll Using Arnon Method • Estimation
of -Carotene Using Spectrophotometric Method
Water Analysis: Introduction • Determination of Dissolved Oxygen in
Water Sample • Determination of Chemical Oxygen Demand in Water
Sample • Determination of Biochemical Oxygen Demand in Water
Sample
Microbiology: Study of the Growth of Aspergillus Niger by
Determination of Inorganic Phosphate and Sucrose Uptake and
Measurement of Citric Acid and Mycelium Produced • Determination of
Inorganic Phosphate and Sucrose Uptake and Citric Acid Production •
Determination of the Amount of Citric Acid in a Plant Tissue •
Growth Curve for Escherichia Coli • Study of Mutation by Ames
Test
Nutrition: Proximate Analysis of Food Material • Sampling •
Proximate Analysis • Vacuum Oven Method • Vacuum Desiccator •
Determination of Crude Fat • Determination of Crude Proteins •
Determination of Ash Content • Straight Ash Combustion Method •
Glycerol – Alcohol Method • Determination of Crude Fiber •
Nitrogen-free Extract • Growth Studies and Determination of Protein
Efficiency Ratio • Group Feeding • Animals • Individual Feeding •
Preparation of Stock Diet • Group Feeding • Nitrogen Balance
Studies on Barley K • Principle • Plan of the Experiment • Material
and Method • Diets • Animals • Metabolic Cages • Feeding • Weighing
• Collection of Feces and Urine • Nitrogen Determination
Chromatography: Introduction • Partition Chromatography •
Requirements for the Partition Chromatography • Separation of Amino
Acids/Sugars by Paper Chromatography • Separation of Amino Acids
Mixture by silica Gel G Thin Layer Chromatography • Separation of
Plant Pigments by Thin Layer Chromatography • Gas (Gas-Liquid)
Chromatography • Basic Equipment • Carrier Gas • Ion Exchange
Chromatography • Determination of the Capacity of the Ion Exchanger
• Isolation of Desired Enzyme by Using DEAE Cellulose Ion Exchange
Chromatogrophy • (A) Regeneration of DEAE Cellulose • (B) Packing
of Column with DEAE Cellulose • (C) Chromatography • Gel Filtration
Chromatography • Gel Matrix • Determination of Molecular Weight of
a Protein Using Sephadex G- • Whitekar Method • Method of Andrews •
Affinity Chromatography • Binding of Ligand on Agarose
Electrophoresis: Introduction • Agarose Gel Electrophoresis (AGE)
for Nucleic Acids • Factors Affecting Migration of DNA through
Agarose Gels • Detection of Nucleic Acids • Polyacrylamide Gel
Electrophoresis (PAGE) for Nucleic Acids • Polyacrylamide Gel
Electrophoresis (PAGE) for Proteins • Significance of SDS-PAGE •
Isoelectrofocusing • Western Blotting • Enzyme-linked Immunosorbent
Assay (ELISA) • Extraction of Nucleic Acid from Agarose and
Polyacrylamide Gels • Mini Agarose Gel Electrophoresis •
Polyacrylamide Gel Electrophoresis for DNA Sequencing •
Immunoelectrophoresis • Rocket Immunoelectrophoresis • Restriction
Fragment Length Polymorphism (A Non-PCR Based Marker) • DNA Probes
• Randomly Amplified Polymorphic DNA (RAPD) • Amplified Fragment
Length Polymorphism (AFLP) • Southern Blotting • Northern
Blotting
Centrifugation: Introduction • Differential Centrifugation •
Density Gradient Centrifugation • Ultracentrifugation •
Determination of Molecular Weight Using Ultracentrifugation
Polymerase Chain Reaction (PCR): Introduction • General/Traditional
PCR • Materials Required •Visualizing Results • Other Important PCR
• Reverse Transcription Polymerase Chain Reaction (RT-PCR) • RT-PCR
Involves Two Steps • A Reverse Transcription • B. Real-time PCR
Plant Tissue Culture Experiments: Laboratory and Equipment • A
Plant Tissue Culture Laboratory • B Equipment Required for a Plant
Tissue Culture Laboratory • Culture Media • Plant Tissue Culture
Media • Sterilization • Sterilization of the Medium • Sterilization
of the Glassware • Sterilization of the Instruments (equipment) •
Sterilization of the Plant Material • Sterilization of the Transfer
Area • Sterilization of the Culture Room • Single Cell Culture •
Isolation of Single Cells • Culture Methods • Batch Culture •
Continuous Culture • Measurement of Cell Growth in Suspension
Cultures • Detection of Viability of Cells in Suspension Cultures •
Cell Synchronization • In vitro Regeneration of Plants • Explants •
Callus Induction • Plant Regeneration • Organogenesis • Factors
Affecting in vitro Regeneration • Micropropagation • Advantages of
Micropropagation • Somatic Embryogenesis • Technique • Importance
of Somatic Embryogenesis • Protoplast: Isolation, Culture and
Fusion • Introduction • Selection Procedures for Fusion Products •
Cybridization • Haploid Production
Appendix I • To Calibrate ml Volumetric Flask • Absolute Error or
Deviation • Mean (average) Deviation • Relative Mean Deviation
(RMD) • Relative Error • Relative Mean Error (RME) • Standard
Deviation (S) • Variance (V) • Coefficient of Variance (CV)
Appendix II • Separation of Micromolecules and Macromolecules Using
Dialysis
Appendix III • Concentration of a Sample Using Ultrafiltration
Appendix IV • Raising Polyclonal Antibodies in Rabbit
Appendix V • Ouchterlony Double Immunodiffusion Technique
Dr. Anil Kumar is Full-time Professor of Biotechnology at the Devi Ahilya University, Indore, India, since 1991. He has also been heading the School of Biotechnology since then. He is a founder member of the School. He has published more than 100 full length research papers in various international and national journals of high repute. He also authored a number of books and contributed many chapters to various edited books. Dr. Sarika Garg is a post-doctoral scientist at the University of Saskatchewan, Canada. Earlier, she worked at the Max Planck Unit for Structural and Molecular Biology, Hamburg, Germany, for more than three years. She has many research papers to her credit. She is the co-author of the book Enzymes and Enzyme Technology. She has co-authored many chapters of various edited books. Ms. Neha Garg has submitted her Ph.D. thesis at the Barkatullah University, Bhopal. Earlier she worked as Scientific Researcher at the University of Potsdam, Germany, for more than three years. She has many research papers to her credit. She is the co-author of the book Genetic Engineering and several chapters in edited books.
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