Escherichia Coli: Purpose Of Bacterial Transformation

This pBlu lab had for purpose to present the changes of the strain of E. coli bacteria due to new genetic information being introduced into the cell. In this experiment we are freezing and heat shocking the E. Coli bacteria that is then forced to take the plasmid DNA. The E. coli then transforms the pBLu plasmid, which carries the genes coding for two identifiable phenotypes. After following the Carolina Biological steps our lab worked well and we able to see some colonies of bacteria on the plates. The x-gal plate showed a significant amount of bacteria to confirm that the pBlu plasmid took over the E. coli strain.

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

To do a transformation one micro test tube was labeled +pGLO and the other was labeled –pGLO, using a transfer pipette 250 µL of transformation solution was added to each tube in the foam rack. The tubes were then placed on ice for three minutes. During these three minutes, a sterile loop was used to pick up a single colony of E. Coli from the starter plate by gently running the loop over the agar. This loop was then inserted into the +pGLO tube and the loop was spun until the entire colony dispersed. Using a different sterile loop, the same procedure was used for the –pGLO tube. After both tubes had their own colony of E. Coli, they were placed on ice for another three minutes. DNA plasmid was added to the +pGLO tube by taking a new sterile loop and immersing it into the stock tube creating a film across the loop then inserting

If a gene that codes for Green Fluorescent Protein transforms bacteria and GFP glows when transformation occurs, then when two micro test tubes have 250 microliters of transformation solution and places in an ice bath, then 2-4 bacteria colonies are added to each tube with a sterile loop; then a plasmid (pGLO) is added to one of the tubes, incubated in ice for 10 minutes, then heat shocked for 50 seconds at 42 degrees Celsius, then back into 9ice for two minutes; then LB nutrient broth is added to both tubes (250 microliters) and set out at room temperature for 10 minutes. Then, 100 microliters of each solution in the tube are added to four

Griffin was a scientist that showed that there was a non-living component to heritability between organisms, although he had not yet discovered what exactly was the heritable component. However, during this time period, it was a common belief that the heritable material was contained within the protein of the organism, not the DNA that is now held in the truth. In the Avery and company experiment, they expanded Griffin research to pinpoint the exact location of the heritable information by breaking down cell components until the final product allowing transformation was

The goal of this experiment was to create a “super e. coli,” as mentioned in the introduction, by co-transforming the pKan and pGlo plasmids. Since the e. coli were not able to absorb both plasmids at the same time, it can be noted that the transformation did not work. The plates containing the pKan and pGlo plasmids separately demonstrated a successful transformation, indicating the bacteria’s inability to take in two plasmids at the same time. Both of these plates acted as our controls, so in the end we could

Coli bacteria, and mix it with the plasmid. We can select the bacteria that are resistant by introducing ampicillin. Bacteria that are not resistant to ampicillin dies. The screening of the bacteria with the resistance occurs when the bla gene turns on to produce beta lactamase which will kill the antibiotic ampicillin. After this, we can get the plasmid into the bacterial cell through the process of horizontal gene transfer through transformation. This can be done by a cold treatment in an ionic solution, then a heat shock where it increases membrane fluidity to take up the plasmid into the bacteria cell. In order to know if the EPAS1-TD gene product was produced by the bacteria, we have to perform a Western Blot. We do this by extracting the protein and run it through gel electrophoresis to separate the proteins by their size. We then label the proteins with the colored probe based on their size. We can detect the EPAS1-TD gene being produced based on the colored

The objective of this experiment was to observe the transformation occurrence with E.coli and the ampicillin resistance gene.When plasmids like lux or pUC18 are added in E.coli, they are more likely to survive in certain environments that contain antibiotics. Plasmid can carry genes enabling bacteria like E.coli to survive in harsh conditions. This experiment displayed how plasmid work when inserted in E.coli with and without ampicillin. In certain agar plates, ampicillin was added with either the lux plasmid or the pUC18 plasmid resulting in colony growth. In other agar plates, no ampicillin was added when either the lux plasmid or the pUC18 plasmid was added resulting in colony growth. If growth occurred from the bacteria when it was added in the Ampicillin agar plates, then it was determined that the E.coli was transformed successfully and is expressing the Ampicillin resistant genes.

According to our lab manual, Genetic transformation is when the genetic makeup of an organism is altered by incorporating external genetic material (Barnhart and Hopper 2014). Genetic transformation was discovered on accident by the scientist Frederic Griffith in 1928. He was trying to create a vaccine for pneumonia, but instead discovered bacterial transformation (Barnhart and Hopper 2014). Plasmids, which are a genetic structure in a cell that can replicate independently of the chromosomes, are used as a form of extra genetic material in order to transform bacteria. They are expressed by both the bacterial cell and the daughter cell. In the case for our genetic transformation lab, our cells we are using are E. coli cells, and

In two weeks of genetic transformation, we were able to successfully complete the objectives of the lab by correctly performing a transformation of bacterial cells. During the lab, we were able to successfully complete the components that were necessary for a transformation of bacterial cells. Viable cells were transformed, able to grow, and were put in a sterile environment conducive to growth. With the exception of the plate labeled LB/Amp: -pGLO, our results followed the lines of a successful experiment. The LB/Amp: -pGLO plate was contaminated and had colony growth in it. In order for the transformation to be completely successful the LB/Amp: -pGLO plate would have had zero colony growth. As stated in the laboratory manual, biotechnologists

the bacterial chromosome is genetically engineered and the plasmid is used to help the bacterium replicate.

There were two parts of this lab and part 1 was; Transforming E. coli with the pGreen plasmid and Part 2; PCR and Electrophoresis. For this lab, a genetic transformation procedure was performed to introduce a plasmid to another cell and when the cell reproduces it will make a new copy of the plasmid. And genetic transformation is a process whereby genetic materials that are carried by individual cells are changed adding foreign DNA into its genome. Also, a“Plasmids are pieces of double-stranded DNA that can be replicated independently of chromosomal DNA, and normally

By using the ligation mixture formed in experiment 3B, competent cells undergo transformation in this experiment. The cells do not usually exist in a state of transformation ready but they can be made permeable to the plasmid DNA and the capable of transformation are referred as competent. Transformation is the uptake of heritable information in a competent cell. Competent cells care extremely fragile and need to be handle gently to ensure the success of transformation. Water bath is heated to 42oC as heat shock treatment to start transformation. The competent cells containing recombinant vectors can be identified through blue white screening. Cells containing recombinant vectors are able to survive on Luria Bertani (LB) medium supplemented with the antibiotic, penicillin. The cells transformed by recombinant vectors are plating on plate that containing isopropyl β-D-thiogalactopyranoside (IPTG) which as an inducer of the lac promoter, and 5-bromo-4-chloro-3-indolyl- β-D-galactoside or called X-Gal which is a dye that produces a blue colour when

Transformation is used in several areas, such as DNA cloning (make multiple copies of DNA) and to produce large amounts of human proteins (insulin). Transformation is the change in genotype and phenotype due to the assimilation of external DNA by a cell (Urry et al. 2016). When naked DNA is incorporated into bacteria’s genetic material, this is when transformation occurs in bacteria. The success rate of genetic transformation is more significant and faster in bacteria, such as E Coli due to the single-celled trait of the bacteria. When these types of experiments are usually performed, a plasmid DNA is used. A plasmid is a small circular DNA molecule that carries accessory genes separate from those of a bacterial chromosome; in DNA

Bacterial transformation is the process of moving genes from a living thing to another with the help of a plasmid.The plasmid is able to help replicate the chromosomes by themselves; laboratories use these to aid in gene multiplication. Bacterial transformation is relevant in everyday lives due to the fact that almost all plasmids carry a bacterial origin of replication and an antibiotic resistance gene(“Addgene: Protocol - How to Do a Bacterial