In this experiment the bacteria E. Coli will be genetically transformed into a competent by going through a process called Heat shock. Heat shock is when you take a bacterial cell and have sudden increase in temperature which increases the permeability of the plasma membrane this causes the cell to take up the DNA from the surrounding medium. (Lab Manual) There are several other methods of genetic transformation but in this lab those will not matter. In this specific experiment pGLO will be the medium around the E. Coli. Genetic transformation is the active up take of foreign DNA in a bacterial cell. (PubMed) For genetic transformation to occur we have to have a medium that contains a different DNA than the thing we are trying to genetically transform. The medium in this experiment will be the plasmid pGLO. This pGLO plasmid is a vector, which transfers a gene from one organism to another. (Lab Manual) The plasmid contains the GFP (green fluorescent protein) gene which makes the bacteria glow in the presence of a sugar called arabinose. The pGLO also contains a gene for resistance to the antibiotic ampicillin. Ampicillin will be used to see if the bacteria lived because the ampicillin is an …show more content…
I predict, that after 24 hours, that the E. Coli that took up the pGlO plasmid will have the GFP gene and with the GFP gene if there is sugar present I believe that you will see the bacteria glow in the dark. Only the cells that take up the pGLO plasmid will survive when ampicillin is present because the ampicillin will kill the bacteria without the pGLO. The results of this experiment are important to know because, you can find out if you had an error in your procedure. Also if you know the results you can sometimes maybe you can leave out some of the experiments you are trying because you know they will not work. We know the results from what we read in the lab
Once the recombinant plasmid was obtained, it was then inserted into E. coli cells through transformation. From a successful transformation, we expected the bacterial cells to translate the inserted EGFP sequence into its protein form. The bacteria cultures were plated on petri dishes containing growth supplement, Luria Broth (LB), an antibiotic: Kanamycin, and IPTG which induced the fluorescence property within successfully transformed bacterial colonies. Different variants of the petri dishes were also included as control and unknown.
Figure 2 shows the results of the electrophoresis. Lanes 5 and 7 indicate the fragments obtained when the plasmids are digested with both restriction enzymes, indicating the approximate fragment size for the hlyA gene, the pK184 plasmid and the pBluescript plasmid. This is useful for identifying the recombinant DNA needed for this experiment
The ligation was expected to make four combinations. The original pBK-CMV and CIH-1 fragments would region to make a non-recombinant pBK-CMV/CIH-1 plasmid. The original pUC19 fragments would rejoin to make a non-recombinant pUC19 plasmid. The larger fragment of pBK-CMV and the small 27bp fragment of pUC19 or the desired recombinant vector, CIH-1 fragment and the larger 2659bp pUC19 fragment. As pBK-CMV does not contain the ampicillin gene then transformed Ecoli containing these would not to survive on the Agar leaving only pUC19 recombinants and non-recombinants.
It has an outer membrane that contains lipopolysaccharides, a periplasmic space with a peptidoglycan layer, and an inner cytoplasmic membrane. It also consists of adhesive fimbriae. Some strains of E. coli are piliated and are capable of accepting, as well as transferring plasmid to and from other bacteria. This enables the bacteria under stressful or bad conditions to survive. Although its structure is simple with only one chromosomal DNA and a plasmid, it can perform complicated metabolism to help maintain its cell division and cell growth. E. coli produce very rapidly; a single microscopic cell can divide to form a visible colony with millions of cells overnight (phschool.com). It is the preferred bacteria in most laboratories because it grows fast and easy, and can obtain energy from a wide variety of sources. Since the birth of molecular cloning, E. coli has been used as a host for introduced DNA sequences (biotechlearn.org.nz). In 1973, Boyer and Cohen showed that two short pieces of DNA could be cut and pasted together, and returned to
Abstract: Bacterial transformation involves the change of genetic composition of bacteria by altering its genetic identity. The pGLO plasmid was ingrained in the E. coli cell, which allows the modified E. coli cell to begin to code for the GFP protein gene and the beta lactamase gene (ampicillin resistance gene). After modifying the bacteria cell, the changes involved with the plasmid were tested on 4 plates, two plates containing the pGLO plasmid (+) were treated with LB nutrient media. One of the LB plates contained arabinose in it, which should fluoresce green under UV light. The other LB plate contained ampicillin. Two other plates which did not contain the pGLO plasmid (-) both had LB media growth, one plate just to show cell growth,
...et light. If the LAA plate glows green under exposure to ultraviolet light, then we can conclude that our unknown insert piece of DNA would be the kan gene. If it does not glow green under exposure to ultraviolet light, then then we streak the colony from our LAA plate onto the LAC plate using a sterile glass spreader. When the LAC plate is dray, we place it upside down in the microfuge rack so that it can be incubated at 37 ºC. Incubation at 37 ºC will allow the transformed bacterial cells to grow. If we see bacterial growth on the LA plate containing chloramphenicol, we can conclude that our unknown insert piece of DNA would be the cat gene, since the cat gene is resistant to chloramphenicol. Afterwards, we then grab the microfuge tube labeled NP and repeat the aforementioned steps shown above pertaining to the LA plates. This would be considered our control.
...n ampicillin resistance, and able to decompose ampicillin, while untransformed gene would perish because of ampicillin damaging the bacteria’s cell wall. Moreover, presence of arabinose operon would promotes the binding of RNA polymerase and the genes for GFP are transcribed, and this would result in the bacteria glowing under the UV light. The result of this experiment confirmed the hypothesis. As explained in result section, the transformed bacteria were alive, while untransformed bacteria were dead. Moreover, one that contained arabinose operon grew under the UV light, while one without arabinose operon did not grow under the UV light, since the GFP was not transformed by the arabinose operon. The sample with -pGLO LB was expected to have lawn, since nothing inhibit the cell from growing. On the other hand, the sample with -pGLO LB/amp had no colonies, since the
A human DNA, in which biologists have identified and isolated the gene of interest using probes or antibodies, will then be chosen. This gene of interest is incorporated into the plasmid cuts. These new plasmids are mixed with, and taken up by bacterial cells under suitable conditions. As these bacterial cells reproduce, the plasmids containing the gene of interest will be copied, and transferred to the bacterial progenies. Genes are segments of chromosomes that code for specific polypeptide or RNA molecules. Plasmids are small loops of DNA separated from bacterial chromosomes, or viral vectors. Restriction enzymes are enzymes that cut DNA at highly specific areas that always contains the same sequence of
Heat transfer from high temperature heated surfaces finds considerable application in engineering. Because of its large number of applications in industries, considerable efforts have been made by researchers to investigate various aspects of the heat transfer and its fundamental principles involved. Fluid flow problems involving heat transfer viz. in presence of convention and radiation represents an idealization of many meaningful problems in engineering practice. Due to the presence of higher level of temperature required in many system like boiler, nuclear reactor; the effect of radiation heat transfer increases. So, there becomes a need of including radiative effect of the participating medium and also their boundary conditions. Keeping this in mind, an attempt was made to investigate the heat transfer in the Indian Pressurized Heavy Water Reactor (IPHWR) during Loss of Coolant Accident (LOCA) with low steam flow. This study will help in estimating the safe working limits for the heat dissipation in the reactor.
Beating the heat is an article about a girl who went on vacation with her family to Arizona. However, one of the worst fire disasters was occurring. Yarnell Hill ignited into flames due to a lightning bolt. Firefighters from the Prescott fire department went to go battle the flames. Using the crew set out to clear brush and trees in the way of the fire. However due to a sudden change in wind the firefighters found themselves trapped by a 100ft wall of fire and 19 members lost their lives.
This process gives you the ability to identify the length and specific genotype of DNA. Due to DNA having a negative charge it was attracted to the positive current provided by the light which made it migrate. The separation of the short molecules from the long ones was obvious because the short ones move much faster when exposed to the positive charge. I used the proper amount of voltage and allowed the gel to process the correct length of time. I used several different times which allowed me to view the migration at a slower pace. The ethidium bromide dye illustrated the migration pattern of my samples. The experiment was a success because I was able to determine the correct genotype for each
The report is written to explain DSC, the thermal analysis technique. In this technique the differential analysis on the base of reference material is done at different temperature. A very close and similar technique is DTA (Differential Thermal Analysis) . In these technique the material is heated at different temperature although sometimes isothermal analysis also done for specific applications. The temperature is recorded for any heat release or absorption. So the heat capacity is measured at those temperatures. Two possible modes for DSC are power compensation mode and heat flux mode DSC. So, DSC is a technique which measure the heat capacity at various temperature of material and reference.
Genetic engineering is a laboratory technique used by scientists to change the DNA of living organisms. DNA is the blueprint for the individuality of an organism. The organism relies upon the information stored in tits DNA for the management of every biochemical process. The life, growth and unique features of the organism depend on its DNA. The segments of DNA, which have been associated with specific features or functions of an organism, are called genes.
Heat energy is transferred through three ways- conduction, convection and radiation. All three are able to transfer heat from one place to another based off of different principles however, are all three are connected by the physics of heat. Let’s start with heat- what exactly is heat? We can understand heat by knowing that “heat is a thermal energy that flows from the warmer areas to the cooler areas, and the thermal energy is the total of all kinetic energies within a given system.” (Soffar, 2015) Now, we can explore the means to which heat is transferred and how each of them occurs. Heat is transferred through conduction at the molecular level and in simple terms, the transfers occurs through physical contact. In conduction, “the substance
Heat Treatment is that the controlled heating and cooling of metals to change their physical and mechanical properties whereas not changing the merchandise kind.Heat treatment is usually done unknowinglyas aresults of manufacturing processes that either heat or cool the metal like attachment and forming. Heats Treatment is sometimes associated with increasing the strength of cloth but it can also be accustomedalterbound manufacture ability objectives like improve machining, improve formability, and restore plasticity when acold operative operation. so it is a awfully sanctioning manufacturing methodology that will notsolelfacilitate completelydifferent completely manufacturing methodology but can also improve product performance by increasing strength or different fascinating characteristics. the heattreatment operation ar oftenoutlined