Pglo Negative Lab Report

The color of the bacteria was a whitish color and the colony size is similar both before and after the transformation. The best way to do it is to compare the control of the experimental plates. Cells that were typically not treated with the plasmid could not grow on ampicillin, although cells that were treated with the plasmid can grow on the LB/AMP plate. The plasmid would have to confer resistance to ampicillin. Moving on, the GFP gene is what is glowing in the plate because it was activated by the sugar arabinose. The sugar arabinose and the plasmid DNA are also needed to be present because that is what initially turns on the GFP gene which makes the bacteria glow. Organisms can also turn on and off particular genes for camouflage reasons. An organism would benefit from turning on and off certain

As predicted the E. coli colony transformed with either the PUC18 or the lux plasmid developed an ampicillin resistance. Which made it easier for them to not only survive but also replicate in both the LB agar plates and the LB ampicillin rich agar plate. However the E. coli colony not treated with the plasmids could not survive and colonize in the LB ampicillin rich agar plates. The plate that had no ampicillin in its environment and no plasmid treated E. coli served as a positive control for this experiment because it demonstrated how the E. coli would colonize and grow in a normal setting. The cells in the positive control plate grew into lawn colonies because they were not placed into a selective environment or transformed, so they had no need to acquire ampicillin resistance. Two plates in the experiment contained E. coli cells that were transformed with either the PUC18 or the lux plasmid but were placed in an ampicillin free environment. These two colonies grew

The plasmid pGLO contains an antibiotic-resistance gene, ampR, and the GFP gene is regulated by the control region of the ara operon. Ampicillin is an antibiotic that kills E. coli, so if E. coli, so if E. coli cells contain the ampicillin-resistance gene, the cells can survive exposure to ampicillin since the ampicillin-resistance gene encodes an enzyme that inactivates the antibiotic. Thus, transformed E. coli cells containing ampicillin-resistance plasmids can easily be selected simply growing the bacteria in the presence of ampicillin-only the transformed cells survive. The ara control region regulates GFP expression by the addition of arabinose, so the GFP gene can be turned on and

Coli cells would engage in the pGLO plasmid and fluoresce. one funding did not support the hypothesis. One plate that contained arabinose did not no glow because the cells did not engage in the pGLO plasmid, due to the lack of ampicillin on that plate. The presence of ampicillin is a motivator for the E. Coli cells to engage in pGLO, which also contains beta-lactamase. According to the student manual, “Beta-lactamase acts against ampicillin, protecting the cell from the antibiotic’s harmful effects. Without the threat of ampicillin, the cells have no reason to take up the pGLO plasmid, and therefore the presence of arabinose has no effect on whether the cells fluoresce or not” (Lab

What was expected that the strains of E. coli that did not have a resistance to ampicillin would not grow. The transformed strain also changed to a blue color when the X-gal was present in plate. The transformed cells also grew because they were free of the ampicillin because they possessed the amp gene that they used as an shield against the ampicillin antibiotic. The transformed cell who turned blue did so because the gene converted the sugar to a blue color but also contained the amp gene to ensure that they grew even when the ampicillin was present. The growth of the colonies on the plates

Next heat shocks the tubes for 50 seconds, followed by icing for 10 more minutes. The heat shock increases the permeability of the cell membrane to DNA. Then add 250 milliliters of LB and incubate for 20 minutes. The 20 minute incubation following the addition of LB broth allows the cells to grow and express the ampicillin resistance protein, beta-lactamase, so that the transformed cells survive the subsequent ampicillin selection plates. Plate 100 microliters the + tubes evenly on two plates; 1 of LB and Amp, and one of LB, AMP, and ARA. Plate 100 microliters of the – tubes evenly on two plates; 1 of LB and AMP, and one on LB only.

4) A MSA plate test was run on a sample of the organism and the results were consistent with the given results for M. Luteus. The results showed a fair amount of growth on the plate, and the color of the agar around the growth remained red. The MSA test is selective in that the salt will inhibit most gram negative organisms and select for gram positives. If there is growth and the color of the agar turns yellow around the growth, this would mean that mannitol was fermented by the organism and the acid waste released by the bacterium lowered the pH around the growth. Since there was growth and no color change, the sample is said to be gram positive and unable to ferment mannitol (negative for differential). This result was also consistent with the given test results for M. Luteus.

The bases of this experiment was to discover the identify of the unknown from three possible specimens: Klebsiella pneumonia, Escherichia coli, and Enterobacter aerogenes. Utilizaing the T streak technique, the bacteria was isolated into pure colonies for further study. The Gram Stain method was used to identity the morhphology of the bacteria such as the shape and whether the bacteria was Gram positive or Gram negative. Biochemical test were also used to help identify the unknown bacteria. The biochemical test used was the Triple Sugar Iron Agar, Sulfur Indole Motility test, Methyl Red test, Voges-Proskauer test, Citrate test, Urease test, and the Gelatin test. After observing the morphology of the bacteria using the Gram Stain method and utilizing all the possible biochemical test, the bacteria was identified to be Enterobacter aerogenes.

The objective of this experiment was to identify two unknown bacteria from a mixed culture. Which was done by using the aseptic technique which was very important to avoid any contamination and keeping the workspace clean while culturing bacteria for different tests. To start, I chose a tube which had a solution of mixed culture. I used the flame to sterilize the inoculating loop and dipped it into the tube and streaked for isolation on 2 TSA plates and placed them in an incubator at 37 for 24 hours. Next day I observed the growth of 2 different types of colonies one for each unknown on the two plates. So I picked the best one and labeled it as master plate and discarded the other plate. From the master plate, I subcultured each type of colony

Similarly, the recipient E. Coli bacteria is resistant to nalidixic acid, and would be able to grow on a Nal plate, but not on the Cm plate. By plating the recipient bacteria on chloramphenicol, we can ensure that the sample was purely recipient if there is no growth.

It takes about 15 days for lab results to be complete once samples are received. The FADL emails lab results to the requestor and will CC you in the email. It is important that you keep track of the lab results; you will need to make a deposition for all lab positive or failed lab results. Notify CW5 Finch or APHC representative when samples contain pathogens. AR 40_657.pdf Chapter 5 section 5-6 give detailed instruction for nonconforming lab samples. When you receive a positive sample lab result that is non-pathogenic, you will make the decision on the which course of action to take. Recently I received lab results for product that had a high SPC counts. I contacted the facility management address the issue, issued the manager

Control #1 would have a known response, where the competent cells transformed to be ampicillin-resistant. In other words, Control #1 should grown on both LB plate and LB+antibiotic plate. Control #1 would provide a good positive response for comparison with the other unknown responses (growth or no growth) of the treatments by different enzymes that broke down each of the three components of the transforming extracts. Therefore, it could be seen whether a component broken down by enzyme was essential for the transformation (transfer and inheritance). Control #2 was the negative control because no response was expected, and Control #2 should not have a response. In other words, Control #2 should grow on the LB plate but not on the LB+antibiotic plate. If it did have a response, there must have been something wrong with the experiment (eg. contamination of the competent cells).

The LB/amp- plasmid petri dish had no growth because ampicillin was incorporated. Ampicillin is an antibiotic that kills potential growth for a bacteria. The LB-Plasmid petri dish eventually grew a lawn of bacteria, this dish only had Laurel broth which is food for bacteria, explaining the substantial growth. We observed circular colonies that only grew on a little corner of the dish. This awkward growth is probably due to the inaccurate spreading of the cells.The LB/amp+plasmid petri dish had growth as well because the ampicillin resistance gene was able to grow since the plasmid contained a gene allowing for antibiotic resistance. The LB+plasmid petri dish had growth because there was no antibiotic to inhibit

A positive control is when you test your experiment against something where you know what the effects will be. a negative control would be when you test the experiment with something you know will have no effect.

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