Molecular Biology & Chromatography Techniques Course

Explore how fermentation uses aerobic and anaerobic microbes to produce metabolites, enzymes, and antibiotics, and examine bacteria, fungi, gut microbiota, and industrial roles in biofertilizers and bioinsecticides.

Molecular biology and fermentation rely on microbes as fast multiplying, carbon-efficient factories whose production depends on regulatory gene-protein-metabolite networks and specific metabolic pathways that steer enzymes like protease and cellulase.

Explore the four metabolic pathways driving microbial growth—glycolysis, the pentose phosphate pathway, the TCA cycle, and the glyoxylate shunt with acetate. Block any pathway; biomass will not increase.

Explore alcoholic fermentation, including mixed acid and 2,3-butanediol pathways that convert glucose to pyruvate, enabling butanol production via Clostridium and propionic acid production via propionate bacteria.

Source industrial microbes from culture banks like NCIM at NCL Pune and apply bioprospecting in environmental niches to screen for metabolite production, including enzymes and antibiotics.

Isolate microbes through primary screening with serial dilution, inoculation on media, incubation, and microscopic/gram-based identification; then perform biochemical tests and secondary screening to assess feedstock carbon sources for product formation.

Explore how feedstock availability, seasonal storage, and transportation costs shape industrial fermentation economics, compare sugarcane and corn bioethanol, and examine feedstock complexity and pretreatment in sustainable production.

Identify process parameters for solid and liquid feedstocks, including particle size and morphology, moisture content, water holding capacity, and biochemical composition. Consider turbidity, viscosity, and pH adjustments as needed.

Form media by balancing inorganic and organic nitrogen sources and minerals; optimize one-at-a-time variations and add precursors, inducers, and elicitors to boost enzyme production.

Use sodium bisulfite to repress acetaldehyde, bromide to curb chloride formation, and penicillin to enhance permeability, while optimizing carbon and nitrogen sources, minerals, vitamins, media, and bioreactor operation.

Explore sterilization in fermentation, including steam sterilization with brief exposure, chemical methods (ethylene oxide, ethanol, sodium hypochlorite), and filtration for heat sensitive materials, guided by Arrhenius kinetics.

Modify media with added protein for protease from rice straw and use orange peel for pectinase. Explain spirulina production in algal ponds, including media, agitation, filtration, drying, and tablet formation.

Screen soil isolates to find microbes producing xylanase, pectinase, and cellulase. Use primary screening with wheat bran, citrus peel, and waste paper; secondary Congo red and iodine tests confirm activity.

Optimize bioprocess economics by balancing media and supplement costs, process runtime, and starter culture viability, while selecting fermentation mode and substrate to maximize yield, titer, and product quality.

Operate batch fermentation as a closed system with fixed media and microbes to maximize substrate utilization and product recovery; primary metabolites form in exponential growth, antibiotics in stationary phase.

Understand fed batch fermentation as a modified batch process, with feeding timed at the end of exponential phase, using fixed or variable volume and intermittent or incremental strategies.

Optimize batch fermentation for products directly linked to microbial growth, yielding high cell density and biomass through an intermittent feeding regime that optimizes baker's yeast biomass and minimizes ethanol.

Explore continuous fermentation, including chemostat, turbidity, and oxo state controls, using biosensors for pH, dissolved oxygen, and CO2 evolution to manage single, recycling, and multi-stage processes.

Compare submerged fermentation with liquid substrates like broth or molasses, including fed-batch and continuous, to solid-state fermentation on water-absorbing solids where moisture and particle size influence diffusion and growth.

Identify substrates: starchy substrates enable amylase; protein-rich oil cakes enable protease; lignocellulosic materials support cellulose activity and lignin degradation; defined media with inert carriers ease purification and may lower costs.

Explore the physico-chemical factors shaping solid-state fermentation, including microorganism selection, inoculum quality, moisture content and aeration, and how pH, temperature, particle size, and porosity influence growth and product yield.

Compare submerged and solid state fermentation for lipids using trichoderma harzianum. Submerged uses olive oil and yeast extract, while solid state with castor bean cake yields lipolytic activity.

Explore production metrics: titer, yield, and productivity that quantify upstream bioreactor performance and downstream purification. Learn how strain improvement and process choices boost titer and yield toward commercial production.

Explore strain improvement through induced mutation using chemical mutagens and radiation, with mutagenesis, selection, and screening to assess enhanced product formation in industrial strains.

Explore how x-ray and uv mutagenesis amplified the A106 kb region in Penicillium chrysogenum, boosting penicillin G production through 5–6 fold gene amplification and increased mRNA and enzyme levels.

Screening and selection drive mutagenesis, varying mutagen concentrations to manage many microbes; the process is tedious and inefficient, with concerns about genetic stability, robustness, and media requirements.

Explore metabolic engineering by engineering metabolic pathways to improve strain production, including heterologous protein production, expanding carbon sources, enabling new product, optimizing cellular physiology, reducing byproducts, and boosting yield.

Compare growth coupled engineering with growth decoupled production engineering to explain how metabolic pathways boost cell density and yield.

Engineers rewire E. coli metabolism to enable co-utilization of glucose with arabinose and xylose by deleting the glucose phosphotransferase system, enabling poly lactate glucose glycolate synthesis.

Explore tolerance engineering to enhance microbial strains' resilience against material toxins, metabolites, and product inhibition in ethanol production, using adaptive laboratory evolution, genomic or transcriptomic engineering, and screening and selection.

Explore how genetic stability and strain robustness enable scalable microbial cell factories, comparing insulin production in E. coli and yeast with mammalian systems.

Learn how recombinant insulin was produced in two E. coli strains expressing A and B chains, then purified and joined by disulfide bonds; compare proinsulin, lispro, and glargine designs.

Explore bioreactors as vessels for microbial growth from small flasks to stainless steel, with automation and sensors ensuring sterility and scalable aerobic or anaerobic production.

Explore bioreactor geometry, including vessel diameter and height, impeller spacing, and baffles, with an aspect ratio of about 1 to 3 to optimize axial mixing, aeration, and heat exchange.

Master impeller geometry for homogeneous mixing, exploring radial and axial flow and blade designs—from flat vertical blades to elephant ear impellers—optimizing oxygen transfer with low shear.

Simultaneous axial and radial flow is achieved by angling flat blades at 45 degrees, enhancing mixing and oxygen transfer with open, semi-open, and closed impellers.

Understand how mixing-induced shear stress damages cell membranes and fungal hyphae, and how controlling speed, impeller type, and anti-foam use maintains headspace and reduces foaming.

The lecture explains fully automated bioreactor control systems, contrasting basic sensor–actuator loops with advanced algorithms that integrate pH, temperature, and oxygen to optimize valve operations and anti-foam responses.

Explore submerged fermentation in a bioreactor, handling energy and carbon sources, anti-foam agents, and nutrient regimes; learn about batch, fed-batch, continuous cultures, chemostat controls, and seed culture optimization.

Master bioprocess parameters by analyzing adequate mixing to improve oxygen transfer, recognize shear forces from mixing that limit growth, and manage submerged fermentation where most bioreactor content is liquid.

Classify bioreactors by agitation into pneumatically driven, mechanically agitated, and non agitated systems; non agitation favors solid state fermentation, while submerged fermentation uses liquid with pneumatic or mechanical agitation.

Explore pneumatically driven gas–liquid bioreactors, including airlift and bubble column types, their riser and downcomer zones, sparging, and suitability for large-scale aerobic production.

Explore orbitally shaken bioreactors for small-scale mixing with gentle stress compared to impellers, and examine pre-sterilized, disposable mixed bag bioreactors using rocking motion for animal and microbial cell culture.

Explore solid state fermentation with no free water, contrasting aeration and mixing strategies across tray, packed bed, and rotary drum bioreactors, including substrate handling and humidity control.

Explore rotating drum bioreactors with no forced aeration, where a horizontal cylinder provides continuous or intermittent mixing and ensures oxygen transfer when the substrate bed is not high.

Contrast SSF with SMF to show how substrate and water content drive bioreactor design, mixing, and online process control.

Illustrates solid state fermentation in a rotary drum bioreactor, detailing seed culture and inoculum handling, rotation-driven mixing, and downstream extraction of the solid substrate.

Explore metabolism in microorganisms, distinguishing anabolism and catabolism, with enzymes regulating energy generation and macromolecule synthesis such as proteins from amino acids.

Explore how enzymes act as biological catalysts, transforming substrates like proteins, starch, fats, and pectin, with proteases such as pepsin and trypsin and plant sources like papain and bromelain.

Learn enzyme types by occurrence, distinguishing intracellular (endo) and extracellular (exo) enzymes, and review production—from cell disruption and fermentation to purification and formulation.

Explore enzyme specificity, including group and absolute types with alkaline phosphatase, glucose oxidase, and glucose isomerase. See how monomeric, oligomeric, and multimeric forms govern activity and inactivation.

Isozymes are enzyme variants that catalyze the same reactions but differ in amino acid composition and physicochemical properties, with examples like hexokinase and lactate dehydrogenase in heart versus skeletal muscle.

Classify enzymes by reaction type, show common and tribal names, and map them to six ec classes and their four-part ec number.

Categorize enzymes by application into technical, food, and animal-feed; technical enzymes dominate detergent, textile, pulp-paper, leather, chemicals; amylases and cellulases lead carbohydrates, with proteases and lipases also used.

Food grade enzymes from generally regarded as safe microbes require certification; dairy uses microbial rennet and acid proteases, while wine employs pectin enzymes to boost juice yield, color, and flavor.

Explore how the starch industry produces high fructose corn syrup from corn starch and apply biocatalytic stain removal using amylases, proteases, lipases, and cellulases in detergents, with cost implications.

Explain how feed enzymes such as phytases break down phytic acid, and proteases improve mineral and amino acid availability, boosting growth and performance in poultry, ruminants, swine, and aquatic species.

Explore therapeutic enzymes with anti-tumor, anti-inflammatory, antioxidant, and digestive applications, and learn why grass organisms are considered for safe industrial enzyme use in the Indian market.

Enzymes bind substrates at a specific active site to form an enzyme-substrate complex and catalyze reactions, releasing products; the video contrasts lock-and-key model and induced-fit model.

Compare the lock and key and induced fit hypotheses, where the lock and key implies a rigid active site, while induced fit shows the active site changing shape upon binding.

Explore how enzyme activity responds to enzyme concentration, substrate concentration, temperature, pH, and inhibitors, including competitive and non-competitive inhibition at allosteric sites, with saturation and denaturation concepts.

Discover how enzymes rely on cofactors - organic coenzymes or inorganic metal ions - forming apo and holoenzymes with prosthetic groups to lower activation energy and define Km and Kcat.

Explore paper and thin layer chromatography to separate components by solubility, using RF values in the mobile phase; the stationary phase guides separation, with detection by ninhydrin or UV.