Hi Olya! Great question. If we know Shweta’s subcluster, we can compare it to other phage within that subcluster. This would be important because if another phage within that subcluster is useful in phage therapies or can produce useful gene products, Shweta may be able to as well.
Hi Brian! We use a program known as DOGEMS, which stands for Deconvolution of Genomes after En Masse Sequencing. Using DOGEMS, we can mix several samples of phage DNA from different clusters together and sequence them all. Because the genomes are different since they are from different clusters, DOGEMS can piece these individual sequences together to create full sequences. DOGEMS allows for the sequencing of several phages rather than just one, so it is ideal for a class such as ours where we isolate several unique phages. Here is a link to a SEA-PHAGE website that has more information of DOGEMS: https://seaphages.org/media/docs/DOGEMSDescription.pdf
Hi Damien, thank you for the question! So Siphoviridae phage have an average tail length of 150nm and head diameter of 55nm. Phage do not need to have these exact measurements to be a Siphoviridae phage. Siphoviridae phage are known to have the longest tails, so any tail length that is longer than the 150nm average would be considered a member of the Siphoviridae family. Myoviridae and podoviridae are other families of phage, but Myoviridae have much shorter tails while podoviriade have even shorter ones. The last family, inoviridae, are known to be worm like and don’t really have a head. This is why Shweta would be considered Siphoviridae.
Great presentation! How would you be able to isolate the phage to treat a patient in the long-run? Also, in general, would you be able to specify the future directions for the long term in addition to short term? Very good explanation of your experiment though.
Hi Mandy, thank you for the question! In our lab we were able to directly isolate our phage by collecting a soil sample and then perform several experiments to purify and isolate a single unique phage, which would be too much detail for this response haha! From there, we can store this sample at 4 degrees celsius or in a refrigerator and the phage would still live. However, we also have designated a portion of our phage lysate to be stored at the University of Pittsburgh and the University of Colorado Boulder so that others have access to our phage. If our phage became of interest for medical reasons, both of these institutions could give out our phage for further testing. As far as future directions go, the short term would be to reperform restriction digest and PCR to ensure that our characterization of Shweta was accurate. In the long term, we would like to sequence and annotate Shweta’s genome so that we can understand its gene functions.
Why is it important to know which subcluster Shweta belongs to?
Hi Olya! Great question. If we know Shweta’s subcluster, we can compare it to other phage within that subcluster. This would be important because if another phage within that subcluster is useful in phage therapies or can produce useful gene products, Shweta may be able to as well.
What method would you use to sequence Shweta’s genome?
Hi Brian! We use a program known as DOGEMS, which stands for Deconvolution of Genomes after En Masse Sequencing. Using DOGEMS, we can mix several samples of phage DNA from different clusters together and sequence them all. Because the genomes are different since they are from different clusters, DOGEMS can piece these individual sequences together to create full sequences. DOGEMS allows for the sequencing of several phages rather than just one, so it is ideal for a class such as ours where we isolate several unique phages. Here is a link to a SEA-PHAGE website that has more information of DOGEMS: https://seaphages.org/media/docs/DOGEMSDescription.pdf
Why is Shweta put in the Siphoviridae mycobacteriophage when Shweta’s has a different length than Siphoviridae mycobacteriophage?
Hi Damien, thank you for the question! So Siphoviridae phage have an average tail length of 150nm and head diameter of 55nm. Phage do not need to have these exact measurements to be a Siphoviridae phage. Siphoviridae phage are known to have the longest tails, so any tail length that is longer than the 150nm average would be considered a member of the Siphoviridae family. Myoviridae and podoviridae are other families of phage, but Myoviridae have much shorter tails while podoviriade have even shorter ones. The last family, inoviridae, are known to be worm like and don’t really have a head. This is why Shweta would be considered Siphoviridae.
Great presentation! How would you be able to isolate the phage to treat a patient in the long-run? Also, in general, would you be able to specify the future directions for the long term in addition to short term? Very good explanation of your experiment though.
Hi Mandy, thank you for the question! In our lab we were able to directly isolate our phage by collecting a soil sample and then perform several experiments to purify and isolate a single unique phage, which would be too much detail for this response haha! From there, we can store this sample at 4 degrees celsius or in a refrigerator and the phage would still live. However, we also have designated a portion of our phage lysate to be stored at the University of Pittsburgh and the University of Colorado Boulder so that others have access to our phage. If our phage became of interest for medical reasons, both of these institutions could give out our phage for further testing. As far as future directions go, the short term would be to reperform restriction digest and PCR to ensure that our characterization of Shweta was accurate. In the long term, we would like to sequence and annotate Shweta’s genome so that we can understand its gene functions.