21 thoughts on “P80 – McCollor

    1. Hi Marika,
      The gel was created in lab for us. It was a 0.8% gel (0.8 g of solid gel agarose in 100 mL of TBE 1X Buffer) which is pretty standard for conducting gel electrophoresis. We unfortunately had to skip making the QC gel due to time constraints within the class. We would not have been able to conduct any restriction digest or electrophoresis experiments had we not skipped it and that was more important to my partner and I than achieving the best possible results since our phage could not be archived.

  1. Can the plaque morphology tell you any other information about your phage? Also does your phage have a name?

    1. Hi Erica,
      This is a great question! The plaque morphology can tell us a lot of information about our phage. Primarily we use it to determine the lifecycle we predict the isolate is undergoing however it can tell us about the concentration of the lysate, whether or not the host cells are a lysogen for the phage that is trying to infect it, and even give us an indication if multiple species of phage are present. Since we were unable to archive our phage, it does not have an “official name” however my partner and I have referred to it as CakePop because of the EM images we obtained.

    1. Hi Kaleigh,
      We had multiple different initial sample sights. Unfortunately after nearly 5 rounds of enrichment we were unsuccessful in obtaining any phage from the soil we collected. This inevitably led us to “adopt” a phage lysate from another group that had success in finding phage. In general the sample location should be somewhere that is easily accessible, contained a sufficient amount of soil, life was present (plants, trees, insects, etc.), and there were no biocontaminents present (chemicals or substances that can potentially harm the environment, affect the life present around the area).

  2. Would the fact that your isolated bacteriophage is lytic make it more applicable in terms of treatment against antibiotic-resistant bacteria, due to their ability to immediately lyse the host cell?

    1. Hi Holly,
      You are exactly correct! For the implications of the newer Phage Therapy technique being researched, it is most beneficial for the phages to be lytic in terms of their abilities to infect and kill the host bacterial cell. If the phage were temperate (able to undergo the lysogenic lifecycle) then it would not be effective in killing the host bacterial cell of interest and thus not be any better at treating the infection as opposed to the antibiotics being administered.

    1. Hi Sarah,
      The enrichment protocol involved incubating the soil sample with laboratory M. Smeg bacteria at 37°C for 48 hours on a shaking incubator and then adding phage buffer to form a lysate. This is what we used to plate our first round of plaque assays to determine whether phage was present in our sample or not. This was one of two isolation methods and proved to be most useful for the research we conducted due to time constraints.

  3. Why do you think that your gel electrophoresis results were inconclusive? If they had been conclusive what outcome would you have expected?

    1. Hi Sarah,
      Our results from the gel electrophoresis experiment were inconclusive likely due to the amount of DNA isolate added to the wells. If we would’ve had more time to conduct a quality control gel, the amounts of the agents used in each well could have been measured more precisely and allowed for more verifiable results. If they had been conclusive, we expected our phage DNA to be cut by at least 3/6 enzymes OR show more cuts for one specific restriction enzyme indicating that our phage belonged to a cluster in which that enzyme has a particular affinity for slicing the DNA.

  4. Thanks for your presentation! How would different cluster results in future experiments affect application of this bacteriophage in a clinical setting?

    1. Hi Zoe,
      Excellent question! Depending on the cluster we could classify this phage into would determine how we could apply this phage in a clinical setting. There is no guarantee that our phage could have implications for Phage therapy or be a candidate for that type of treatment however, by classifying our phage to a cluster we can then try and further characterize it through sequencing methods and other bioinformatic tools. This in turn allows for the diversification of the field of phage biology and hopefully one day will be useful for treating antibiotic resistant bacterial infections once the technique has been perfected and approved for clinical application.

  5. Thank you for your presentation! How does being a lytic or temperate phage affect applications in a clinical/medical setting?

    1. Hi Zoe,
      I touched on this topic in a previous question above. Lytic phages are the phages of primary interest when it comes to clinical applications for their ability to infect and kill the host cells. This pertains mostly to bacterial infections that antibiotics cannot treat and thus is the reason for the newly sparked interest in developing techniques to use them in clinical treatments.

    1. Hi Bryton,
      The results from the gel electrophoresis experiment were not conclusive enough for us to characterize our phage to a particular cluster or grouping. This does not mean however that it does not have a group that it belongs to. Our results unfortunately did not provide us with enough evidence to be able to make that classification and assign it. I hope that cleared up any confusion!

  6. Fantastic presentation. What potential does this classification of phage have for future directions?

    1. Hi Kyle,
      The classification of our phage would aid in it’s characterization. This would allow us to conduct some sequencing and bioinformatic experiments to better understand the gene products our phage produces and compare them to other known phages in either similar or different clusters. This then opens the door for a variety of genomic and potentially clinical applications once characterized.

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