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Lecture 3

Laboratory Exercise - Week 3.pdf

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Biological Sciences
Daman Bawa

Brunt 2011 BIOB12 Laboratory 3 Microbiology Module Growth and enumeration of EcoliOutline Lab 3A monitoring of Ecoli growth using both optical density and viable plate counts Lab 3B analysis of Ecoli growth data and the preparation of LB plates containing the antibiotic ampicillin for lab 4 1 Objective to develop the skills needed to grow and monitor bacterial growth is the foundation technique for most molecular biology experiments requiring recombinant DNA moleculesLearn to work with Escherichia coli E coliLearn aseptic techniqueProper disposal techniques for bacterial culturesMaster concept of dilution growth curve construction andcalculatinggeneration time 2 Note you are working with live bacterial culture follow all procedures covered byyour TA and report any spills immediately If you are unsure of any step ask Introduction toBacterial growthBacteria are a diverse groupof simple prokaryotic cells In BIOC17 you will have an opportunity to experience the great diversity of these organisms For the purposes of a molecular biologist the workhorse eubacterium is Escherichia coli E coli In nature Ecoliis a natural inhabitant of your gut It server a beneficial roleCertainnatural stains of Ecoli howeverare pathogenic andcan make you sick Molecular biologists however do not use wild type Ecoli Molecular biologists use mutant strainsspecifically designed to optimize the uptake and expression of foreign DNAby E coliThese mutant strainsor laboratory strains cannot grow outside the laboratoryCan you think of a reason why this is very important There are over 100 different molecular biology strains available that differ in their genotype and researches therefore can choose the appropriate strain for their needs E coli is the most commonly used bacterium in cell and molecular biology labs for propagation of recombinant DNA molecules In order to use these strains researchers must be able to properly grow and monitor the growth ofthese bacteria In this laboratory we will monitor growth of E coli by two different methods referred to direct and indirect methodsE coli can grow on a carbon source like glucose supplemented withsimple inorganic salts However the more nutrient rich the media the faster the bacteria grow In molecular biology we use rich media to grow bacteria Growth of a microorganisms is defined as an increase in cell numberFor most molecular biologyexperiments E coli is grown in arich medium known as LB Luria Broth or LuriaBertani broth You made bothLB 1
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