The purpose of this experiment is to make E.Coli competent so that it can be transformed in order to become immune to ampicillin, then we would be able to determine the transformation efficiency of the culture. We determine this by preparing 4 plates of E.coli, each labeled “LB-plasmid”, “LB+plasmid”, “LB?Amp-plasmid”, and “LB/Amp+plasmid”. This meant that either should have lacked plasmid and Ampicillin, with plasmid but lacked Ampicillin, without plasmid but with Ampicillin, or were with Ampicillin and plasmid, respectively. Then we made the bacterial cells competent by adding CaCl2 to 2 vials of the colony (one with plasmids), and incubating on ice, then heat shocking, and returning to ice. Luria Broth is then added and left to sit for 5-15
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
Escherichia coli, or better known as E. Coli, is one of the most commonly studied single-cell organisms because it is easier to manipulate and it is abundant. Some E. Coli strains can be pathogenic, but most are harmless. They can help benefit digestive health by filling niches in place of more harmful microorganisms. Because it is a diverse group of related microorganisms, it can be difficult to find the boundaries defining the species. Some strains are natural while others are genetically created in labs, which can be observed in aspects of an organism (Schussler).
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
E. coli HB101 was transformed with pGLO plasmid then grown on media containing ampicillin and/or arabinose and on medium containing neither (Brown, 2011). This is done for selection of transformed cells since not all cells are expected to take up the plasmid (Brown, 2011). We also expect roughly the same CFU on any plate(s) receiving samples from the same microcentrifuge tube, since they are getting the exact same
I inoculated a T-Soy agar with bacteria number 118, for this I used a streak isolation method. Next, in order to distinguish between Gram positive and Gram negative I used a streak isolation technique on a CNA plate, then performed the same exact procedure on a MacConkey plate. The results from the CNA plate showed the Gram Positive bacteria was an Alpha hemolyzer. Next, I used a P Disc on a T-Soy agar inoculated with bacteria 118 and determined the Gram Positive bacteria was not sensitive to P Disc antibiotics. This revealed the Gram Positive bacteria to be Streptococcus Mitis. The results from the MacConkey plate proved the Gram Negative bacteria to be a lactose fermenter. With the Gram Negative bacteria I performed a lysine test with positive results. Next, I performed an ornithine test on the Gram Negative bacteria, with negative results, therefore I concluded the Gram Negative bacteria was Klebsiella pneumoniae.
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
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.
E. coli is a bacterium capable of being transmitted from one person to another. To ensure that the bacteria used in this experiment were not transmitted to other places, items, or organisms, several safety protocols were followed.
CDC says that 63 people were infected with the outbreak strains of E.Coli 0121 and that 026 have been reported from 24 states. The sickness started from December 21, 2015 to September 5, 2016 and 17 sick people were hospitalized. CDC found out that some people who got sick had to of eaten or dealt with raw dough. FDA’s investigation determined that the raw dough restaurants were using General Mills flour that had to of been produced in November of 2015. General Mills flour manufactured that it was likely to be the source of the outbreak. General Mills contacted all of the costumers who have gotten flour from there and told them about the recall. FDA and General Mills are working together to make sure that the costumers have all been notified.
In order to test the stability of the bead in the different temperature, the bacteria activity was measure by using pour plate count method. This method is to show the stability of Escherichia while store in freezer at (-4 0C) and incubator (37 0C) for 1 week. Data collects everyday and total plate method was apply to count the amount of bacteria for immobilization process.
During this report the demonstration will establish an awareness to the audience, which discovers the statics and findings of the results based on samples from a mathematical strategy perception. This possibly identifies the conclusion of the significant evidence of exposure with the amount and levels of unwanted consumption of E. coli in the Village of Sierra Leon’s drinking water. During the 5 weeks the missionaries have to achieve safer drinking water for the village, through three types of sampling using one hundred for the first and second tests, and twenty for the final evaluation. The population discovery consists of various elements such as the mean, median, and mode. That offers presentation for the average center tendencies.
The specimen was processed similarly to case-1. Escherichia coli was grown in aerobic culture, and Bifidobacterium sp. was cultured in anaerobic culture. The identification of Bifidobacterium sp. was done by both MALDI-TOF Vitek MS and Vitek-2. Escherichia coli was found to be Extended spectrum beta-lactamase (ESBL) positive and sensitive to Piperacillin+Tazobactam, Cefoperazone+Sulbactam, Imipenem, Meropenem, Amikacin, Gentamicin, Tobramycin, Chloramphenicol, and Cotrimoxazole. Bifidobacterium sp. was found to be sensitive to Penicillin, Ceftriaxone, Imipenem, Meropenem, Amoxycillin+clavulanic acid, Piperacillin+Tazobactam and Clindamycin and resistant to Metronidazole. The patient showed a good response to Meropenem and recovered completely.
In this lab experiment, students had to create a growth curve for E. coli. The E. coli growth curve would illustrate the progression of the population of E. coli a set time period. In this case, the growth curve depicted the population of E. coli over a 12-hour period. The growth curve for E. coli was created from the absorbance levels, the optical density(OD), recorded from the spectrophotometer.
Life on this planet began with microorganisms. Through millions of years microorganisms have found ways to successfully adapt and survive. These adaptations have created a wide biodiversity, allowing them to basically populate in all places. Why are these microbes so important? Because they shape the history of our world. Some microbes can be deathly to humans while some others are favorable, for example, bacteria that lives in the gut of both humans and animals and helps during the process of digestion (Alfred Brown & Heidi Smith, 2006). Understanding these interactions help scientists to find ways to protect humans from potential deathly pathogens. In order to observe microbes, microscope proficiency and microorganisms’ identification are crucial skills in a microbiology lab. During this laboratory session, samples of environmental and human organisms were inoculated into two different rich media and incubated to their according temperature. After this, appropriate use and calibration of the microscope was performed. Lastly, morphology and size of different species of bacteria, algae, fungi and protozoan were recorded.
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.