Lab Exam

To adress the growing problem of antibiotic resistance by the discovery of new antibiotics through student research

Select a area and roughly collect 5 - 10 grams of soil in tube. Mark down date and location of soil collection.

Serial dilutions are needed to ensure that there is a countable number of colonies on plate, allowing for accurate estimate during colony counting

Start with a sample and add 1 gram of the sample to larger ammount of dilution. From that add an ammount of the sample into the first dilution tube. Now the original sample doesnt get used anymore and

Targets eukaryotic cells so they don't grow on media. Inhibits growth of unwanted bacteria.

5.2x10^7 rather than 52,000,000 or 3.2x10^-5 instead of 0.000032

Colony Forming Unit/ Per gram of soil

# of coloniesx dilution factor/ volume of dilution

30-300 are ideal

Luria broth (LB plate) because it is non selective medium allowing for a wide range of bacteria and Trypticase soy agar (TSA plate) because it is also non selective and allows for a wide range of bacteria to grow

Acts as a standard reference point to assess the performance of a test due to its predictable behavior in various environments

This is a group of 6 extremely antibiotic-resistant bacteria such as Staph aerus, and Klebsiella

Safe relatives are those closely related to highly antibiotic-resistant pathogens but are considered non pathogenic making them useful for research purposes such as staph epidermis, E. Coli, Enterobacter aerogenes

Master plates contain isolated colonies of bacteria and allow for transfer to other plates and allow you to keep track

Take the bacteria from the master place and place the bacteria on 2 new plates within a grid formation and place the tester organism

Narrow would show resistance against one or the other tester strains and broad would show resistance againt both E. coli and staph

The purpose is to isolate individual bacterial colonies from a mixed sample

Gram Stain
Endospore stain
Nutrient agar
Motility agar
Fermentation tubes
Indole test
Methyl red
Voges proskaeur
Citrate
Hydrogen sulfide
Catalase
Blood agar
Crystal violet agar
EMB agar
Manitol salt plate

Antibiotics are more effective against gram-positive bacteria because gram-positive lacks the outer membrane present in gram-negative bacteria. Antibiotics that are more effective against gram negative typically target the outer membrane of bacteria utilizing molecules that can penetrate through porin chanels

We use ethyl acetate because it has polar and nonpolar characteristics the antibiotic is most likely non polar and, because of this should stay suspended in ethyl acetate and the agar and proteins will settle in the water.

The purpose is to isolate the antibiotic from the bacteria

Purified extract was tested for antibiotic activity by taking antibiotic and adding 100ul of methanol to the vial. Then take the vial and place 10ul in one spot and repeating this process 3 times in the same spot allowing it to dry inbetween each placement. We then took top agar and inoculated it with 100ul of tester strain. When everything dries pour the top agar onto spotted media plate. let incubate and assess for zones of inhibition.

To overcome antibitoic resistance by discovering new antibiotics