Week 11

Today's Topics:
○ Biotechnology

Announcements:
○ Exam: 16/1/07

Lecture Topic:

○ What is it? The use of microorganisms, usually on a large scale, to produce valuable products (eg beer, cheese, antibiotics)
○ These are usually enhancements of natural reactions, amplified for better yield.
○ Genetic manipulation now permits production of products that microorganisms do not naturally produce.
○ Main organisms used:
§ Fungi (yeasts & moulds)
§ Streptomyces spp.
§ Escherichia coli
§ Bacillus subtilis
○ Microorganisms are often altered by mutation or recombination, to achieve high metabolic specialisation.
○ Requirements:
§ Must be able to grow on large scale
§ Should grow rapidly, and yield product in a short time
§ Must be able to grow on cheap culture, ideally waste from another process.
§ Should be amenable to GM.
§ Must not be pathogenic
○ Examples:
§ Antibiotics
§ Vitamins
§ Enzymes
§ Alcohol
§ Vinegar
§ Bioconversion
○ Primary Metabolites
§ Produced during growth of culture (eg ethanol)
§ Result of growth
○ Secondary Metabolites
§ Produced at end of growth - at, near, or in, stationary phase.
§ Survival mechanism (eg antibiotics, to kill off competing organisms, or enzymes to free up more food)
§ Not essential for growth & reproduction
§ Dependent on growth conditions (especially medium)
§ Often possible to get massive overproduction.
○ Vessels used are called fermentors, and often hold hundreds of thousands of litres.
○ Two classes: ærobic and anærobic.
○ Antibiotics:
§ Substance produced by microorganisms that kill or inhibit others.
§ Most useful antibiotics produced by filamentous fungi and by Actinomycete group (eg Streptomyces spp.)
○ Penicillin
§ Original: Penicillium notatum - low yield (1-10μgcm^-3)
§ 1943: Penicillium chrysogenum, further improved
§ Now Penicillium chrysogenum, in aerobic stirred fermentors (50,000μgcm^-3)
○ β-lactam antibiotics: Penicillin & its relatives.
§ Biosynthetic penicillins: result of biosynthesis
§ Semisynthetic penicillins: chemically modified
§ Interfere with cell wall production.
○ Vitamins & amino acids
§ Most made by chemical synthesis
§ A few are too complex for this, eg B12 (Propionibacterium spp. & Pseudomonas spp.), and riboflavin (Ashbya gossypii)
§ Amino acids are regulated by inhibitory feedback - mutant species resist this inhibition, and thus overproduce. (Eg use of S-aminoethylcysteine to produce resistant species, where lysine will not inhibit production by aspartokinase).
○ Extracellular enzymes (excreted into medium)
§ Capable of digesting e.g. cellulose & starch
§ Useful in food, pharmaceutical & textile industries
§ Proteases and lipases from alkaliphilic Bacillus lichenformis used in detergents (e.g. biological washing powders)
§ Amylases and glucoamylases: production of glucose from starch.
○ Microbial bioconversion - biotransformation
§ Used for specific reactions beyond organic chemistry
§ E.g. Rhizopus nigricans converts progesterone to 11α-hydroxyprogesterone, which is then converted chemically to hydroxycortisone and then cortisone.
○ Genetically engineered microorganisms
§ E.g. use of E. coli to produce human growth hormone & insulin
§ Contain gene from another organism
§ Grow rapidly on simple media
§ Produce massive amounts of blood proteins, hormones, etc.
§ Main hosts: Escherichia coli, Bacillus subtilis, Saccharomyces cerevisiae.
§ Work from mRNA (to avoid introns), and use reverse transcriptase to make DNA, and then ligate this into a plasmid.
○ Vaccines
§ Killed or attenuated pathogens (or fractions) that produce immunity.
§ Body develops antibodies against surface proteins
§ Genetically engineered antigens
□ Cheaper
□ Easier to purify
□ Free from other proteins
§ E.g. Hepatitis B antigen cloned into S. cerevisiae
○ BT toxin
§ Ulcerates gut wall in larvae
§ Gene transferred to P. flourescens, which inhabits rhizosphere. Innoculation protects roots from pests.
§ Then transferred to plants
§ Constant exposure stimulated BT resistant pests.
○ Herbicide resistance:
§ E.g. gene for resistance to glyphosate transferred into plants, to allow glyphosate to be used to suppress pests without affecting crop.

Colony type 1 - LM400

Colony type 1 - LM1000

Food Culture Plates - observations made 28/11/06

Culture from Salad, on MacConkey Agar, at 1E-3


Culture from Salad, at 1E-4


Culture from Salad, at 1E-5


Culture from Salad, at 1E-5 - culture surface of agar.


Culture from Salad, at 1E-6. I identified 3 cultures on this plate -

Colony type 1 was circular, raised, papillate, red, with an entire edge, and about 4mm in diameter. Under the microscope, it had strings of gram positive bacilli


Colony type 2 was irregular, flat, matt, pink, with an entire edge, and about 15mm in diameter. Under the microscope, it had individual, gram negative bacilli.

Both of these colonies tested positive for catalase, and negative for oxidase.

Colony type 3 was circular, convex, shiny, red, with an entire edge, and about 2mm in diameter.