E. Coli Lab Report

134). They are loops of DNA that are separate from the chromosomal DNA and can self-replicate in a cell, found mostly in bacteria (Brown, 2011; Addgene, 2015). Lederberg and William Hayes discovered that plasmids were being transferred from one cell to another, not the chromosomal DNA (Brown, 2011, p. 135). This discovery lead to plasmids being an essential tool for scientists. Scientists can engineer plasmids to have specific genes to introduce into new cells (Brown, 2011, p. 134). On a plasmid loop there will be an origin of replication (ORI) and a multiple cloning site (MCS) where the gene of interest is inserted (Bio-Rad, 2015). This region has specific restriction enzyme recognition sites, which are cut by the enzymes to open up the DNA where the new gene will be inserted (Jove Science Education Database, 2015). Most plasmids will also contain an antibiotic resistance gene allowing cell survival in environments containing antibiotics (Jove Science Education Database, 2015).

This experiment was designed to test and observe the transformation efficacy of the pUC18 and lux plasmids in making E. coli resistant to ampicillin. Both plasmids code for ampicillin resistance, however, the lux plasmid codes for a bioluminescence gene that is expressed if properly introduced into the bacteria’s genome. The E. coli cultures were mixed with a calcium chloride solution and then heat shocked, allowing the plasmids to enter the bacteria and assimilate into the bacterial DNA. The plasmids and the bacteria were then mixed in different test tubes and then evenly spread onto petri dishes using a bacterial spreader, heating the spreader between each sample to make sure there is no cross contamination. Each of the dishes was labeled and then incubated for a period of 24 hours. The results were rather odd because every single one of the samples grew. Several errors could have occurred here, cross contamination or possibly an error in preparation as every single sample in the class grew, meaning all samples of the bacteria transformed and became ampicillin resistant.

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

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

Escherichia coli O157:H7 causes 73,000 illnesses in the United States annually. A review of E. coli O157 outbreaks reported to Centers for Disease Control and Prevention (CDC) to better understand its epidemiology. E. coli O157 outbreaks revealed that in that period, 49 states reported 350 outbreaks, representing 8,598 cases, 1,493 (17%) hospitalizations, 354 (4%) hemolytic uremic syndrome cases, and 40 (0.5%) deaths. Clinical laboratories began examining more stool specimens for E. coli O157. In 1994, E. coli O157 became a nationally notifiable infection, and by 2000, reporting was mandatory in 48 states. Escherichia coli O157:H7 was first recognized as a pathogen in 1982 during an outbreak investigation of hemorrhagic colitis.

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.

According to a study conducted in a Tunisian Hospital, E.coli resistance rate to ampicillin was 72.1% (Ferjani et al., 2014). Furthermore, penicillin was found to be least effective against E.coli with a 100% resistance rate during a research done in trying to comprehend the patterns of antibiotic susceptibility of bacteria (Saba et al., 2014). E.coli found in the intestinal contents of pigs was found to be 57.7% resistant to streptomycin (Lee et al., 2014). Situated on these past research and studies, if a tryptic soy agar plate in applied with E.coli and separated into four different quadrants with three quadrants having an antibiotic disk and one having the control disk, then the results will show E.coli to be most resistant to penicillin, somewhat resistant to ampicillin and least resistant to

Escherichia coli is a gram negative bacteria which is the main aerobic commensal bacteria found in the intestines of animals; it can also become pathogenic if it gets into the urinary tract ot the blood (Erb et. al., 2007). Staphylococcus aureus is a gram positive bacteria. It is the leading cause of bacterial infections in humans involving the skin, blood, and lower respiratory tract in America (De Leo et. al., 2009); Natural selection of antibiotic resistance in S. aureus has led to the emergence of increased resistance to penicillin, methicillin, and vancomycin within the human population (De Leo et. al., 2009). One important difference between gram-positive and gram-negative bacteria is the structure of their cell walls. The cell walls

It turns out microbes don't wait for five seconds, or even 1 second to hop on to your food. Even brief contact with a contaminated floor will contaminate food wet or dry! Bacteria will adhere to food almost immediately, but time does matter. The food left on the floor for five seconds acquired a lot of bacteria, but the food left on the floor for 30 seconds picked up a few colonies more. Imagine if you left it for a full minute, or even 10 minutes... the amount of bacteria would be ten times greater! Furthermore, don't think your food is safe because the floor "looks clean" or the food that you dropped on it does. Floors make great homes for bacteria. Also, floors come in contact with our shoes on a regular basis, and the University of Arizona

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.

Bacteria has always been one of the most important organisms in the field of molecular genetics, according to scientists. More specifically, Escherichia coli remains one the most famous types of bacteria due to the general harm it causes on hosts. E.coli lives in the intestines of humans and animals, and even though it does not trigger any problems most of the time, certain strains can cause food poisoning. What makes those types of bacteria even more interesting from a research point view, but dangerous when talking about their effect on humans, is the ability to develop antibiotic resistance. Bacteria in general, and E.coli in particular, have small circular DNA molecules called plasmids, which are separate from the bacterial chromosome. Those plasmids carry one or a few genes and usually encode for proteins that serve as a protection from one or more antibiotic. A particular strain of E.coli carries the plasmid pAMP. This particular plasmid confers resistance to ampicillin, a penicillin-like antibiotic.

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.

Antibiotics have been used to treat patients who suffer from infectious diseases. However, these drugs have been used for so long that the infectious bacteria managed to adapt to the drugs making them less effective. Nowadays, antibiotic resistant bacteria have become increasingly more common causing the death of thousands of people around the world. Indeed, bacteria possess plasmids. Plasmids are circular DNA molecules, separate from the main chromosome that are replicated and transmitted to daughter cells during cell division. Plasmids can allow the bacteria to live under environmental stresses. Antibiotic resistant genes are found in plasmids. In fact, bacteria with plasmids may have genes that allow them to live under harsh conditions making them antibiotic resistant or immune to some toxins. Thus plasmids carry and spread resistant antibiotic gene in a short amount of time through cell division in wild population.

A gene must be transferred from one plasmid to another in order to give that organism a selective advantage e.g. antibiotic resistance. However, Bacteria have been industrialized that are resistant to antibiotics such as penicillin which is a result of the misuse of antibiotics. Resistance arises as changes or mutations within the bacteria’s genes develop, which help bacteria grow that are resistant to antibiotics and therefore have an advantage. From researching journals, I found that Pseudomonas Aeruginosa resistance poses threats to human and animal health as the body no longer has the defences to protect against the bacteria. Altogether 77 isolates were tested and out of 77, 13 where negative for the presence of class 2 integrase gene (Olga et al, 2016). Only the most beneficial bacteria are selected to carry plasmids and so natural section is an important factor so that only those of the greatest advantage are selected. There are various different types of plasmids some of which include F-plasmids for expression of the sex pilli, resistant plasmid that provides resistance against antibiotics or poisons, col plasmid which contain proteins that kill or

The excessive and frequent use of antibiotics in medical treatment largely contributed to the development of antibiotic resistance (Sengupta, 2013). But, bacteria that have never been exposed to antibiotics also exhibit antibiotic resistance showing that it is an ancient and natural phenomenon (Bhullar et al, 2012). Antibiotic resistance can result due to intrinsic qualities of the bacteria or it can be acquired through genetic methods such as mutations or horizontal gene transfer through plasmids. Antibiotics kill susceptible bacteria but are ineffective against resistant bacteria, this creates selective pressure, which allows the resistant bacteria to grow and multiply. This leads to the development of highly resistant pathogenic