By getting an accurate cloistering for your phage as a future direction, how will this benefit your research? Will you be able to compare it to other clusters to further progress your research/question?
By getting an accurate cluster of my phage this will make future use of my phage a lot easier. If someone were to use it in phage research or medical treatment experimentation and we had an accurate cluster of the phage as well as the sequenced whole genome associated with the clustering then they would be able to seek out specific DNA information that would go well with their research. As of right now we can compare only the morphology to other phages but not clusters since we did not get as far as sequencing our phages DNA genome.
Bacteriophages are categorized into related clusters based on the comparative analysis of their whole genome sequences. By analyzing the whole genome sequence, one is able to accurately cluster these phages with other phages that have related genomes. We never got to this point in our experimentation.
Some specific methods and techniques used to obtain the phage were enrichment of a soil sample which involved getting dirt from outside and adding a Lysogeny Broth media and Mycobacterium smegmatis mc2155 to it in order to obtain the solution that contained the phage. From then on there was a lot of serial dilutions of the solutions to purify and isolate the phage itself. Other than that the figured describe techniques used to get the very final results.
Classifying and clustering my phage would contribute to the future and be helpful by making future use of my phage a lot easier. If someone were to use it in phage research or medical treatment experimentation and we had an accurate cluster of the phage as well as the sequenced whole genome associated with the clustering then they would be able to seek out specific DNA information that would go well with their research.
It is kind of hard to prevent denaturing of the DNA because we do not know it is denatured until we look at the gel. Denaturing can be cause by the DNA solution being heated. The heating causes the double-stranded DNA to unwind and the hydrogen bonds that hold the two strands together weaken and they eventually break. At first we believed that we might have accidentally left our DNA solution out at room temperature too long and that made it denature. The second time we did a gel we took precautions to make sure it was cold constantly but the gel still showed denaturing. So we really don’t know how to prevent it at this point.
You mention that “The calculation of our titer was necessary in order to actually obtain the electron micrograph of the phage in identifying its morphology.” What did this calculation look like? How was it applied/how did it help you obtain an EM image?”
We do show the calculation of the titer in figure 2. It includes the equation we used to calculate the titer as well as the titer calculation total itself. The exact line from the poster is ” Titer (pfu/mL) = 203/ ((10) x (1000μL/1mL) x (10^14). The final titer was: Titer = 2.03 x 10^18.” Obtaining the titer helped us get the EM image because it allowed us to determine the concentration of our phage within our lysate which allowed us to understand what we were looking at in the image itself.
There is really one indicator that shows evident denaturing. This is the blur of the bars looking like they are smeared downwards. If you look at the DNA ladder in figure 3 you see that it shows solid bars that almost form a ladder. The two samples have that blurred smearing downwards and that is what the denaturing looks like.
By getting an accurate cloistering for your phage as a future direction, how will this benefit your research? Will you be able to compare it to other clusters to further progress your research/question?
By getting an accurate cluster of my phage this will make future use of my phage a lot easier. If someone were to use it in phage research or medical treatment experimentation and we had an accurate cluster of the phage as well as the sequenced whole genome associated with the clustering then they would be able to seek out specific DNA information that would go well with their research. As of right now we can compare only the morphology to other phages but not clusters since we did not get as far as sequencing our phages DNA genome.
What is considered to be an accurate clustering of the phage?
Bacteriophages are categorized into related clusters based on the comparative analysis of their whole genome sequences. By analyzing the whole genome sequence, one is able to accurately cluster these phages with other phages that have related genomes. We never got to this point in our experimentation.
What were some specific methods/techniques used to obtain your data?
Some specific methods and techniques used to obtain the phage were enrichment of a soil sample which involved getting dirt from outside and adding a Lysogeny Broth media and Mycobacterium smegmatis mc2155 to it in order to obtain the solution that contained the phage. From then on there was a lot of serial dilutions of the solutions to purify and isolate the phage itself. Other than that the figured describe techniques used to get the very final results.
How does classifying your phage contribute to its in the future? Why would it be helpful to be classified?
Classifying and clustering my phage would contribute to the future and be helpful by making future use of my phage a lot easier. If someone were to use it in phage research or medical treatment experimentation and we had an accurate cluster of the phage as well as the sequenced whole genome associated with the clustering then they would be able to seek out specific DNA information that would go well with their research.
If you wanted to perform another Gel, what would you change to prevent possible denaturing?
It is kind of hard to prevent denaturing of the DNA because we do not know it is denatured until we look at the gel. Denaturing can be cause by the DNA solution being heated. The heating causes the double-stranded DNA to unwind and the hydrogen bonds that hold the two strands together weaken and they eventually break. At first we believed that we might have accidentally left our DNA solution out at room temperature too long and that made it denature. The second time we did a gel we took precautions to make sure it was cold constantly but the gel still showed denaturing. So we really don’t know how to prevent it at this point.
You mention that “The calculation of our titer was necessary in order to actually obtain the electron micrograph of the phage in identifying its morphology.” What did this calculation look like? How was it applied/how did it help you obtain an EM image?”
We do show the calculation of the titer in figure 2. It includes the equation we used to calculate the titer as well as the titer calculation total itself. The exact line from the poster is ” Titer (pfu/mL) = 203/ ((10) x (1000μL/1mL) x (10^14). The final titer was: Titer = 2.03 x 10^18.” Obtaining the titer helped us get the EM image because it allowed us to determine the concentration of our phage within our lysate which allowed us to understand what we were looking at in the image itself.
What are some common indicators in figure 3 that show “evident denaturation”?
There is really one indicator that shows evident denaturing. This is the blur of the bars looking like they are smeared downwards. If you look at the DNA ladder in figure 3 you see that it shows solid bars that almost form a ladder. The two samples have that blurred smearing downwards and that is what the denaturing looks like.
What made you choose this?
Choose what? I do not understand this question.