Discussion 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
Introduction: 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
In 1928, Fred Griffith first discovered genetic transformation by infecting mice with unencapsulated and non-pathogenic pneumococci (Lacks 2003). This was the start that opened up the field of biotechnology. Genetic transformation is a process where foreign DNA crosses a membrane of another cell and then alters the genetic material (Encyclopædia Britannica 2015). Genetic transformation can occur in a few different ways: projectile bombardment, electroporation, and heat shock (Weedman 2015). Heat shock is defined by increasing the temperature of cells environment making the plasma membrane become more permeability allowing new DNA to transfer into the cell (Weedman 2015). The cell receiving the new DNA, also known as competent cells, can either amplify the DNA or clone the DNA (JoVE 2015). The most common type of DNA used to perform genetic transformations is plasmids: small, round DNA molecules that still contains two strands of DNA that has the ability
Transformation Lab Report Introduction Transformation is the transfers of virulence from one cell to another, through the transferring of genetic material. It was originally postulated in 1928 through the works of Federick Griffith, a British microbiologist. Griffith observed that the mutant form, non-virulent form, of the bacteria Streptococcus Pnumoniae could be transformed into the normal, virulent form, when injected into mice along with heat killed normal forms. He concluded that somehow the information the dead virulent form had transformed the mutant form into a virulent form.
Lara Guvelioglu BI 108 E2 Nahomie Rodriguez-Sastre 04/13/17 Bacteria Escherichia coli’s Genetic Transformation Using Vector Plasmid DNA pGLO Abstract The field of biotechnology involves the concept of genetic engineering, altering the DNA/genetic material of an organism using information from a different one. The process in which bacteria can obtain this manipulated genetic information from another source is called genetic transformation. The goal of this experiment was to genetically transform Escherichia coli bacteria’s DNA by inserting the vector pGLO plasmid which codes for ampicillin resistance as well as the green fluorescent protein, GFP. For the experiment, the E. coli bacteria were separated into two groups; control and
Next the tubes were placed in an ice bath, while obtaining a sterile loop to swipe a single colony of E.coli to put into the tube. After gently swiping a colony onto the loop, it was then spun in the +pGLO tube to get it to come off, returned to the ice bath. Next using a different sterile loop, it was swooped it in a container labeled pGLO plasmid DNA and again spun it ONLY into the tube with the solution labeled +pGLO to get it to come off. After about 10 minutes on ice the tubes were then placed into a 42ºC water bath for 50 seconds exactly, and immediately after placed them back into the ice bath. Finally, after 2 more minutes in the ice bath the tubes were separated into 4 containers. 250 ul of +pGLO solution was added to the containers containing +pGLO, LB broth, with ampicillin and +pGLO, LB broth, ampicillin, with arabinose. Also 250 ul of the –pGLO solution was added to the 2 containers containing LB broth, and ampicillin, with LB broth. Using a sterile loop for each plate the solutions were spread out gently and thoroughly on to the containers with agar. After the containers were incubated in 37ºC for at least 24 hours, the results were observed and disposed of (Weedman,
The reason why it did not inhabit any change after the experiment was conducted is because it had no plasmids added to support the growth and the ampicillin actually killed the bacteria. The plasmid would have served as the vector to insert the gene into the E. coli. The -pGLO LB plate thrived because there was no ampicillin present. While the +pGLO LB/amp/ara plate ended up glowing under the UV light because the GFP was only able to show with the presence of the arabinose, because it is what activates the operon. The +pglo LB/amp plate contained a few colonies but it did not glow under the UV light due to the absence of the
After that, a new sterile loop was used to immerse the pGLO plasmid DNA stock tube into the +pGLO tube but no into the – pGLO tube. Both tubes were well closed and put back on the ice for 10 minutes. While both tubes were sitting on ice, the four plates were labeled. LB/amp plate was labeled +pGLO, LB/amp/ara plates were labeled +pGLO, LB/amp plate was labeled – pGLO, and LB plate was labeled – pGLO. After that, the tubes were transferred into a water bath set at 42° C for exactly 50 seconds, then placed rapidly back on the ice for another two
The transformation of E. coli using plasmid DNA was a success. The positive control plate had a near lawn of blue colonies growing on the plate. This indicated that the E. coli cells took up the plasmid and became ampicillin resistant. The blue colonies formed because
Hypothesis: 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
Genetic transformation occurs when an organism’s genetic makeup is altered due to the introduction of new genetic information which is then incorporated into the organism’s genome. In this lab the pGLO plasmid is introduced into E. Coli bacteria, and incorporates the genes which code for the GFP and beta lactamase to the bacteria’s genome which as a result will be modified. To test the effects of the plasmid, bacteria treated with the plasmid were grown on separate plates, the first containing LB nutrient broth and ampicillin, another containing LB nutrient broth and arabinose and another containing LB nutrient broth, ampicillin and arabinose. Two more plates were grown, one with LB nutrient broth and ampicillin and the other with only the LB broth, using cells that did not contain the plasmid. Since the lab was about genetic transformation, the goal was to find which plate would glow. It was found that the plates that were not exposed to the plasmid did not glow, and the plates containing LB and arabinose and LB, ampicillin and arabinose did glow. The plates containing ampicillin, the antibiotic that kills E. coli did not grow whereas the remaining plates at least had some growth.
Mikaylla Holmes Joshua Sanders Niasia Ellis The Bacterial Transformation of pGLO Purpose: The purpose of this experiment is to teach the students step by step on how the genetic transformation process works using pGLO. Genetic transformation is when an individual cell’s genetic material is changed by exogenous DNA.
The objective of this experiment was to genetically transform E.coli cells to express ampicillin resistance and to produce the green fluorescent protein by using the pGLO plasmid. It was hypothesized that only the cells with pGLO DNA added to the solution would be able to survive in the same environment
The Transformation Occurrence of Plasmid in Bacterial Growth Cherline Riche ________________________________________________________________________ Panther ID: 5471556 Crystal Garcia, Joselyn Pozo, Tory Jarett Biology I Lab 1010--Lab Section U55 Abstract: 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 the results of the experiment located on Image 1 in the results section, the phenotype of the E Coli demonstrated a change when exposed to the plasmid pGlo. This conclusion disproved the hypothesis. All four agar plates were exposed to LB broth (Luria broth). This broth is the source of food and nutrients E Coli used to grow and divide. As demonstrated by Image 1, there was growth in the in the agar plates -LB, +LB/amp, and +LB/amp/arab. The substantial growth that was demonstrated in the -LB agar plate (estimated at 75-80% growth) was due to this was the standard E Coli without any ampicillin or pGlo present. Moreover, the agar plate for +LB/amp showed a margin of growth. The percentage of growth was estimated at a 45-50% growth and signified this in individual colonies. This plate contained the plasmid pGlo. This plasmid is coded with GFP protein, antibiotic resistance gene, and the araC gene (www.bio-rad.com). Even though the agar plate contained the antibiotic ampicillin which kills the E Coli bacteria, the E Coli still grew because the plasmid pGLO was present. This growth demonstrated that the E Coli bacteria absorbed the pGLO plasmid and revealed a change in the E Coli’s phenotype. There was a margin of growth because the E Coli was subjected to the transformation solution (CaCl2) and the heat shock. A