Microorganisms In biotechnology

Microorganisms & history Throughout history man has been afflicted by disease. The impact of these diseases have changed the course of history: Malaria- eukaryotic protozoan-Plasmodium. Contributed to the death of King Tut ˜3,000 years ago and the decline of the Roman Empire˜1,500 years ago. Anthrax- Bacillus anthracus – source of the fifth plague of Egypt mentioned in the Book of Exodus in the Bible. Plague- Yersinia pestis– source of the Black Death that killed millions of people during the Middle Aged and resulted in the loss of 30-60% of Europe’s population.

Model Systems In the 1870s German Physicist Robert Koch was able to further studies performed by Davaine and Rayer who used Antrhax as a model system in studying livestock diseases that infect humans. Koch discovered that the anthrax disease was caused by anthrax endospores that persist in the soil. Model systems are useful for gaining knowledge that can be applied to other systems. In the 1880s French chemist Louis Pasteur used weakened forms of anthrax to immunize sheep. Pastuer used what he learned from anthrax to develop a vaccine against rabies.

Koch’s Postulates Koch & Pasteur are recognized as two founders of microbiological research. Koch used his knowledge from anthrax and TB research to develop postulates to determine if a microorganism is the cause of a disease. Postulates: 1. The microorganism must be found in abundance in all organisms suffering from the disease but should not be found in healthy organisms. 2. The microorganism must be isolated from a diseased organism and grown in pure culture. 3. The cultured organism should cause disease when introduced into a healthy organism. 4. The microorganism must be re-isolated from the inoculated diseased experimental host and identified as being identical to the original causative agent. Theses postulates were responsible for the discovery of: diphtheria, pneumonia, meningitis, leprosy, plague and syphilis.

Uses of Bacteria in Biotechnology Food Production Yogurt- lactic acid fermentation Lactobacillus delbrueckii subspecies bulgaricus Protein Production A protein that is artificially produced in a genetically engineered organism is called a recombinant protein. To produce a recombinant protein in bacteria, the gene for the protein is cloned and transferred into the bacteria, it is then cultured which means they are placed in conditions that enable them to multiply into billions of bacteria. Once the culture has reached the appropriate density the bacteria are harvested and lysed. The recombinant proteins are then purified from bacteria using chromatography. Uses for recombinant proteins include therapeutic drugs, agriculture and food production.

Insulin Insulin used to treat diabetes is manufactured as a recombinant protein. Historically purified from pig pancreas. Patient reactions and expense of purification 1978 Genentech cloned the human insulin gene and genetically engineered E coli bacteria to produce human insulin Eli Lilly liscenced and sold under the name Humulin in 1982 Humulin is now produced in Eukaryotic cells

LAB SKILLS TO DATE Follow laboratory protocols Make dilutions from concentrated solutions Use an adjustable volume micropipet Make molar solutions using a volumetric flask Use a serological pipet with a pump or aid Use and calibrate a pH meter Use a transfer pipet Label agar plates and broth Perform calculations C1V1=C2V2 Make LB, LB/AMP, LB/AMP/ARA agar and broth Wash glassware properly Use aseptic technique Make percent solutions Write an SOP Write a label for a reagent bottle Use a spectrophotometer Use a graduated cylinder Calculate molarity Use a balance

Upcoming Lab Skills Inoculate an agar plate for a bacterial lawn Analyze an agarose gel Streak bacteria to isolate single colonies Transform bacteria by calcium chloride transformation Aseptically inoculate a plate with liquid culture Perform HIC chromatography Aseptically inoculate a liquid culture from a plate Perform SDS- polyacrylamide gel electrophoresis Identify bacteria from cell shape Stain Polyacrylamide Gels Prepare agarose gels Dry polyacrylamide gels Load an agarose gel Perform a qualitative ELISA Perform agarose gel electrophoresis Use a power supply