Loop Dilution Lab Report

The purpose of this experiment is to investigate the effects of various household products may have on bacterial growth. The bacteria that was chosen as a test subject is Micrococcus luteus, which is generally considered a harmless bacterium unless the host has a weakened immune system. Among the household products that a set of groups will test are common cleaning and disinfecting products such as rubbing alcohol, Betadine, Germ-X, Neosporin, liquid hand soap, and Lysol cleaner. In the experiment, the bacterium, Micrococcus luteus, is to be swabbed and grown in a Petri dish divided into four sections with agar in each. After that, a small circle of thin paper is soaked in the four household items. Then, a piece of parafilm

solids and liquids that were in the solution. The centrifuge was able to do this

The third procedure we did, our first step was to gather materials of a spot plate,and 2 pipettes one filled with sodium hydroxide and one filled with copper nitrate. We then

(Biology Dept.). 0.1 ml of E.coli K or 0.1ml of E.coli B was added to the 10 fold dilution. Using soft agar technique, the growth media mixture with E.coli was plated and incubated.

70µL of competent E.coli are added to both test tubes; pUC18 and Lux (Alberte et al., 2012). Both test tubes are then tapped and placed back into the ice bath for 15 minutes. While waiting, another test tube is obtained, filled with 35µL of competent cells and labeled NP for no plasmid. A water bath is preheated to 37 degrees Celsius and all three labeled test tubes are inserted into the bath for five minutes (Alberte et al., 2012). Using a sterile pipet 300µL of nutrient broth are inserted into both the control and Lux test tubes and 150µL are inserted to the no plasmid test tube to increase bacterial growth. All three test tubes are then incubated at 37 degrees for 45 minutes. Six agar plates are obtained and labeled to correspond each test tube, three of the plates contain ampicillin. A pipet is used to remove 130µl from each test tube containing a plasmid and insert it into the corresponding agar plate. For this, a cell spreader is first

14. Use the same loop and technique to spread the same cell suspension (+) on the LB+AMP agar plates. Dispose of the sterile loop in a beaker of germicide.

There were two tubes used in this process: the tube that contained the primary culture and the tube that contained the nutrient agar where the unknown bacteria would grow. First, the inoculating loop was flamed. After removing the caps of both the test tubes, they were flamed to prevent contamination of the unknown bacteria. The inoculating loop was cooled for a few seconds and was then placed into the test tube containing the bacteria. The inoculating loop with the bacteria was placed into the nutrient agar test tube for cultivation. Before the test tubes were capped, they were flamed once again. Also, isolation of the unknown bacteria had to completed. Nutrient agar was placed in the petri dish, and was left to gel for a few minutes. After the agar gelled, the inoculating loop was used to acquire bacteria and streak the unknown onto the plate for

Spin the two tubes in a centrifuge for 5 minutes on opposite side of the centrifuge. The bacterium will collect at the bottom of the tube, so pour out the extraneous supinate. Then, add 250 microliters of buffer. The Ca2+ cation of the buffer neutralizes the repulsive negative charges of the phosphate backbone of the DNA and the phospholipids of the cell membrane allowing the DNA to pass through the cell wall and enter the cells. Place both tubes on ice. Then add 10 microliters of water into one tube and 10 microliters of plasmid DNA into another tube labeling the one with DNA with a + and the one with water -, and place on ice for 10 minutes.

coli bacteria affects bacterial growth and gene expression in several environmental conditions. To begin, two micro test tubes were labeled; one with +pGLO and one with -pGLO. 250 µl of transformation solution was added to each microtube, with use of a sterile pipette. Afterwards, the pipette was placed in a bleach solution and the microtubes were placed into the foam rack and onto crushed ice. Next, a colony of bacteria was removed from the starter plate with a sterile loop and thoroughly mixed within the +pGLO microtube. This step was repeated with a second sterile loop and the -pGLO microtube. After their use, all loops were placed in the bleach solution for decontamination and the microtubes were returned to the foam rack and ice.

The test tube was labeled with the bacteria identifying number. The cap on test tube was removed, and the lip of the test tube was flamed. Next, the Bunsen burner was used to sterilize the inoculating loop. Then, bacteria were picked up from the working plate with the loop and the agar was inoculated. The loop was then re-flamed. Finally, the plate was placed in the 37˚C incubator and left to sit for 48 hours and any changes in color were observed. A negative result appears green, and a positive result appears blue. This is because it tests for the organism’s ability to use citrate as its sole source of carbon, and if it does then it produces ammonia and ammonium hydroxide which make the medium basic, changing the green agar blue (Leboffe & Pierce,

In the first part of the experiment, we plated donor bacteria, which was chloramphenicol resistant, on a Nal plate. Because no donor bacteria would grow on a Nal plate, this was a way of ensuring that the sample of donor E. Coli bacteria was pure.

Being able to control bacterial growth is something that is important in our everyday lives. As shown in the previous labs, bacteria can grow and create colonies extremely quickly especially in the right environments. By acknowledging this, it is then important to get an understanding of how bacterial growth can be controlled by humans. To control microorganisms it means to inhibit their growth (static) and or kill them (cidal) (Kenneth Todar, 2015); therefore since focusing on bacteria the terms bactericidal and bacteriostatic are both extremely important for this lab. One broad method we will use to control bacterial growth is heat. The amount of heat needed to control bacterial growth is different for different species of bacteria (Kenneth Todar, 2015). Bacteria can also respond differently depending if moist heating method such as an autoclave with steam is used, or a dry heating method such as inoculating a loop over a fire is used (Kenneth Todar, 2015). UV works by damaging the cells DNA, without proper DNA, the cells will die and the object

This experiment was performed to test the hypothesis if LB nutrient broth, +pGLO and -pGLO Ampicillin, and Arabinose was placed in the E. coli plates, then there will be a significant growth in the newly transformed bacteria and it will possess the ability to glow under UV light. The measurements were recorded from the bent glass tube in each glass test tube. The transformation protocol tested for the newly possessed traits in E.coli bacteria. Throughout the experiment there were many probable reasons for failure. If the pipettes and sterile loop were not thrown out in between each use, a cross contamination could cause a miscalculation in the experiment causing the data results to fail. The hypothesis that was tested was validated due to the positive results with each experiment stating that newly transformed organisms due in fact pass on traits.

According to Encyclopedia Britannica, a microbiologist is someone who studies “microorganisms, or microbes, a diverse group of minute, simple life forms that include bacteria, archaea, algae, fungi, protozoa and viruses” (Pg. 1). On a day to day basis, an average microbiologist will deal with treating infections; identifying and classifying new organisms as they are discovered; and finding new uses for already discovered organisms.

A fresh pipette was used to transfer 0.5ml of broth culture of E. coli, to be inoculated, into a tube of molten agar previously boiled to drive off oxygen. The tube was then rolled to distribute the bacteria and allowed time for the agar to harden. The tube was incubated at 37oC for 48 hours. The procedure was repeated for broth cultures of the bacterium Clostridium sporogenes and B. Subtilis.