Micro Liters Lab Report

In this case, our plasmid DNA is isolated from a liquid culture of the E.coli that was transformed. This is done by reacting the liquid culture with five buffers. The buffers are P1, P2, N3, PE, and an elution buffer. P1 is used to re-suspend the pellet and degrade RNA, P2 is used to lyse cell membrane, PE is used to wash the sample, the elution buffer is used to release DNA from the spin column, and N3 is used to precipitate proteins and genomic DNA. The main components of P1 are Tris, EDTA, and RNase. The main components of P2 are NaOH and SDS. The main component of PE is ethanol and the main component of N3 is acetic acid. The main component elution buffer is water. The possible contaminations of mini-prep are proteins and salts (Garey et al.,

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

It was also conducted to learn about the process of moving genes from one organism to another with the help of a plasmid. The control group was the -pGlo with LB and ampicillin antibiotic, and the -pGlo with LB. The experimental group was the +pGlo with LB and ampicillin antibiotic, and the +pGlo with LB, ampicillin antibiotic, and arabinose. The dependent variable was the bacteria. It had the ability to change. The independent variable is the ampicillin antibiotic and pGlo. They stand alone and are not changed by other factors. The organism used was the bacteria GIVE TYPE OF BAC. and HOW WILL THE BACTERIA BE

The data that my group obtained during this experiment was almost identical to my hypothesis.  On the -DNA LB plate there was bacteria growth because there was no ampicillin to keep the bacteria from reproducing.  On the -DNA LB/amp plate there was no growth because of the ampicillin that kept the bacteria from growing into colonies. The +DNA LB/amp plate had about 234 colonies of bacteria. Not all of the bacteria on the plate were able to grow into colonies but some did because they interacted/picked up the plasmid. Finally, on the +DNA LB/amp/ara transformation plate most, if not all of the bacteria were transformed and, additionally there was also arabinose sugar which activated the araC gene that codes for the green glowing protein which made the bacteria glow up when we shined the UV-light

coli culture incubated at 37 ᵒC and the other one for an E. coli culture incubated at 25 ᵒC. To start the lab, one milliliter of the culture, Luria broth, is transferred to a spectrophotometer tube and zeroed. One milliliter of each culture is transferred to a clean cuvette and the optical density at a wavelength of 595 nm is measured. The OD values and times should be recorded. If the OD value exceeds 1.0, one hundred microliters of the culture and nine hundreds microliters of the nutrient broth must be taken and the OD should be measured again. One of the measured cultures is then transferred into an Eppendorf tube.

In this experiment, the gram negative bacterium Escherichia coli is being subjected to various environmental factors that affect the rate of growth. These factors scrutinized were the different types of nutrients, the intensity of aeration, or the temperature at which it was stored. The purpose of this lab is to determine which factor affects the Escherichia coli the greatest. It is known that these abiotic factors affect the rate of growth the greatest if they remain at the correct conditions for living.

Experiments introduce the idea that the uptake of DNA by competent bacteria can be the result for survival reasons such as need for food or evolution purposes (5). The difference between natural and artificial gene transformation in a bacteria, however, with those who are incompetent versus competent allow he or she to conduct an experiment with artificial transformation. By doing this one may be able to determine and observe the formation of biofilms or mutations by transferring a gene to another organism for beneficial or harmful purpose. This could lead to finding resistants or if a gene introduced has any effect on the bacteria. In the experiment, the pGLO plasmid contained encodes the gene for GFP as well as a gene for resistance to an antibiotic allowing a transformation to take place when adding a carbohydrate such as arabinose to the medium. Research has been found on the horizontal gene transfer on how virulence factors are acquired as well as genes resistant to antibiotics spread with microorganisms (5). By conducting a similar experiment he or she can observe similar findings or observe the results and conduct some sort of understanding of these ideas. The findings in this experiment are to show a positive result in what causes growth and the glowing when the plasmid pGLO is added to the positive as well as the difference in the medium. Understanding the way artificial

The purpose of this experiment was to study the transfer of genetic information on plasmid F’lac by using Escherichia coli. Plasmid transfer was measured by using two different methods. The first one was by using selection and contraselection with three antibiotics: streptomycin(which was replaced by naladixic acid for the second part of the experiment),ampicillin and kanamycin and the second one by using a colour indicator ( X-gal). As significant results, the percentage of transfer for F’lac was higher than the percentage for transposition. Also, the experiment demonstrated that E.coli can quickly acquire resistance to several different antibiotics through the transfer of the F’lac plasmid. It was concluded that significant

(Matthysse et al. 1996) The heat shock protein, E. coli is preserved as both prokaryotes and eukaryotes, however when using the GFP gene, eukaryotes work best. (Lai et al. 2014, Matthysse et al. 1996) The GFP causes the bacteria to glow in the dark, however in order for the fluorescent to happen the gene must be “turned on” in which there must be a presence of a sugar called arabinose. (Sun et al. 2010) The vector, pGLO plasmid, carries the GFP gene and a gene for resistance to the antibiotic ampicillin. The gene resistance to the antibiotic ampicillin is carried by the vector so transformed cell can be screened and by planting cells in ampicillin the only ones that survive by taking up the plasmid will grow and express the GFP gene. Therefore, I hypothesis that in the experiment, the only LB plate that will glow will be the plate labeled +pGLO LB, amp, ara, because it contains all of the necessary components that will activate the GFP gene which will make it

The magnitude of a star is simply how bright the star is. The brightest stars were of first magnitude, however, the dullest were of sixth magnitude. The simple scale of magnitude was made popular by Ptolemy, but is thought to have started with Hipparchus. The scale is sometimes considered confusing since the brighter the star is, the smaller the magnitude it has.

Once the mixture in each tube was completed the opening of the tubes were covered with Parafilm then flipped upside down to completely mix the continent of the tubes. After the solutions were mixed each tube was then wiped down thoroughly using Kimwipes to remove fingerprints and smudges, this is done because imperfections on the outside of the tubes can cause less light to pass through the test tubes causing the lower readings of transmittance that might actually be. Once the test tubes are cleaned the calibration tube is immediately placed into the sample holder (Parafilm must be removed from all of the test tubes before being placed in the spectrophotometer and will be replaced on the tube once the tube is removed from the machine). Then the lid of the spectrophotometer is closed and the calibration button is pressed (this button only needs to be pressed when the machine is being calibrated but the spectrophotometer must be calibrated

Plate three was hypothesized to have little to no growth, due to the fact that it was a pVIB-plasmid negative E. coli strain exposed to an ampicillin positive environment. While this plate did show the least amount of

The bacteria that was used in the experiment was Escherichia coli which also has the ability to communicate through quorum sensing. Quorum sensing controls bioluminescent, antibiotic production, competence this better understand the bacteria that we were working with (Bassler, 2012). We wanted the bacteria to express antibiotic resistance, bioluminescent and growth all of that is control by quorum sensing which is a type of communication between the populations of the Escherichia coli. In addition, Escherichia coli is a gram negative bacteria which appear as purple when is stained. Gram negative have a thin layer of peptidoglycan but the outer membrane has porins which lets antibiotics such as ampicillin to move through the wall and the bacteria can become resistant to antibiotics. In the experiment, one of the way we made the Escherilia coli was make them competent to allow them the express and share the important information between themselves in the form of genes in the selected plasmid. Plasmids help the bacteria to transfer important date for its survival and its one of the features of the plasmids that is used in the transformation. The Escherilia coli first needed to become competent to take the DNA with the help of calcium chloride

Plasmid DNA is negative and bacterium’s cell wall is negatively charged. Thus, as DNA approaches the cell wall it is repelled. The calcium chloride neutralizes the charge of the bacterium and DNA therefore allowing the Bacterium cell wall to be in close proximity to a plasmid DNA. The bacterium undergoes heat shock which is technique of place a bacterium in extreme temperature with wide range. In the lab the bacteria was place in an

Plasmids are small circular DNA molecules. They are not essential for survival of the host bacteria. Some carry genes that allow resistance to antibiotics (Anderson). Plasmid pUC18 is a circular DNA molecule. It contains portions of the E. coli Lac Z gene, which encodes for the first 146 amino acids of β - galactosidase. E. coli contains the Lac Z gene, which encodes β - galactosidase . The E. coli Lac operon digests lactose. Once E. coli is transformed with pUC18, complementation occurs. E. coli produces active β –galactoidase. The active β –galactoidase hydrolyzes the substrate, X gal , which is located on the agar plates.