Mccarty And Macleod: Identifying The Transforming Principle

solids and liquids that were in the solution. The centrifuge was able to do this

The overall goal is to either discover a new novel phage that has not yet been discovered, or provide new research on an already discovered phage. The isolation of the phage takes several steps under the streak protocol that is present in materials and methods section of this report.

5 drops of chloroform were added to the mixture and incubated at room temperature for 15 minutes to kill, and lyse the bacteria cells which allows all the unadsorbed phages to be accounted for when determining the titer of unadsorbed phage with susceptible bacteria, E. Coli B. (Biology Dept.). 0.1 ml of E.coli B was added to the 10 fold dilution. Using soft agar technique, the unadsorbed phage were plated. After incubation, the titer of unadsorbed phage was determined by counting the number of plaques on the plate from the infection of single E. coli cells in the assay tube by a free phage particle before

6 petri dishes were labeled, 3 petri dishes P 105 B, P 106 B, P 107 B, and the other 3 petri dishes P 10 K, P 102 K, P 103 K and they were set aside. A series of dilutions for the 1x bacteriophage T4 rII

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Bacteriophages were first discovered about 100 years ago by Frederick Twort and Felix D'Herelle. Bacteriophages or commonly known as “phages” are viruses that infect bacterial hosts. Bacteriophages come in multiple shapes and sizes. But a good amount of them are tailed viruses that contain double stranded DNA. The head of the bacteriophages has a protein shell which is attached to the tail. Some phages differ as some don’t have double stranded DNA they may be single stranded. Phages are incapable of reproducing on their own. In fact, they need a bacterial host to reproduce. Like any virus, bacteriophages are very fixed to discrete hosts. The host bacteria that we are using is called arthrobacter. Arthrobacter is a genus of bacteria found in soil (Pope et al 2016). Arthrobacter can help reduce hexavalent chromium which can cause some symptoms such as irritations to humans. Hexavalent chromium may also cause lung cancer if inhaled. The SEA PHAGE project was broken down into two parts. The first part which we are currently doing is isolating, purifying and amplifying the bacteria from our soil samples that we collected in the beginning of the year.

The main objectives of this experiment included making dilutions of solutions, plating phage or bacteria, and determining the number of bacterial viruses or phage in a suspension. It was also conducted to demonstrate that two different mutants of phage T4 can exchange genetic material to give rise to wild-type phage. The experiment was used to distinguish mutants from wild-type by their host specificity. The recombination in bacteriophage was performed to determine the concentration of unadsorbed phage from the U series plates, total concentration from B series, and concentration of

This project is all about isolating bacteriophage in soil. They come in different sizes and shapes, each to their own unique look. Phages have a protective protein head that contains DNA and a hollow tube tails (http://phages.org/bacteriophage/). Since bacteriophage cannot reproduce and replicate themselves, they need a host to do the work for

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.

How do you insert the plasmid inside the bacteria? What process do you use? (1/2 pt)

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

This process uses enzymes to cut and insert piece of DNA into a plasmid vector. This vector is then transfected into cells. The different pieces of DNA that were being inserted into this plasmid were a control and three different mutants (an alpha-5-tailless mutant, full-length alpha-5, and full-length alpha-2). These pieces of DNA code for integrins. Once these pieces of DNA were inserted into the plasmid vector, the plasmid needed to be inserted into a cell where it would be able to replicate. In these tests, rat intestinal epithelial wild-type cells (cells that would be found in nature, not cells grown in the laboratory) were used. To insert the plasmid into these cells lipofectamine plus was used. Lipofectamine plus acts like a detergent and opens the membrane of the cell so the plasmid can enter. Also, the plasmids that were transfected into cells were all treated with G-418, an antibiotic. When the cells were plated and allowed to multiply, only those containing the plasmid with the antibiotic would live. This made it possible to know which cells actually received the plasmid and properly underwent mitosis, and to eliminate those cells that never took up the plasmid in the first place.

For this experiment, we have to prepare our phosphorylase which extracted from a potato. We prepared by weighed about 250 grams of peeled potato and cut it into cubes. The extracts then blended with 100mL of 0.1M NaF. After filtered the contents into a clean 250mL centrifuge bottle, we centrifuged it for 3 minutes. Then, separated the supernatant into a centrifuge bottle, which is our phosphorylase preparation. The enzyme assay used in this experiment today is the iodine test. As the iodine reacts with starch, it will form a brown, blue or black precipitate due to the iodine ions forcing into a linear arrangement. The endpoint of the enzyme reaction indicates the presence of starch by using the iodine test to determine. The faster the endpoint is reached, the less active the phosphorylase is.

Stage one the DNA in the pathogens is first transcribed into RNA so that is can be used, as a single strand is needed. Then the pathogens are lysed to expose the nucleic acids and a specific oligonucleotide hydrolyses to the

The petri dish lid was raised to insert the loop. The loop was touched to the agar area on the opposite side of the dish. The bacteria on the loop were transferred to the agar. The bacteria were spread in the first sector of the petri dish by moving the loop back and forth across the dish (zigzag motion).