ISOLATION OF INDIVIDUAL BACTERIAL COLONIES ON SOLID MEDIA
Robert Koch developed a method for isolating pure cultures on solid media in 1883. To this end he added agar (a solidifying agent) to liquid nutrient broth; the nutrient broth supports the growth of a wide variety of microorganisms while the agar provides a solid substrate on which bacteria can be mechanically diluted and therefore isolated as independent colonies representing different bacterial species. The isolation of independent bacterial species from various environmental sources is important in all branches of microbiology since bacteria are ubiquitous and live in microbial communities of mixed populations. Populations in microbial communities or ecosystems may interact and cooperate in their efforts to obtain nutrients from the environment with the waste products from one group of microorganisms serving as nutrients for another. Alternatively, the metabolic wastes from fermentation processes of some bacteria in a population (for example acid production) may provide a favorable ecological niche for bacteria that prefer to grow at low pH. This exercise is designed to teach you how to use solid media and streak plate techniques to isolate pure bacterial colonies from a mixed population of bacteria. The instructor will provide TSA plates (Tryptic Soy Broth mixed with 15% agar, heated to sterilize media and melt agar then poured into a Petri dish) and a culture containing Escherichia coli and/or Bacillus subtilis and/or Staphylococcus aureus and/or Serratia marcescens.
Isolation of bacteria using the streak plate technique:
Label the bottom of a Petri dish (side containing the agar) (Why?) with your initials, date and lab section. Your instructor will provide you with a test tube containing two or more bacterial species. Mix the contents in the test tube by gently shaking the test tube. This will ensure that all of the bacteria are in suspension. Aseptically remove some of the culture...
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