Ladies and Gentlemen, let me introduce you to Golddigger!
Now I’m sure your asking, Why is your phage named Golddigger? If you’ve read my previous blog “The side of scientific research that they don’t tell you about…” then you’ll understand the issues and stress that we had during this years phage hunt and so Golddigger is rather fitting as this phage has demanded alot of my time and energy!
Golddigger was found in compost in the Hobsonville Point community garden in Auckland, New Zealand (as seen on this map; Golddigger was found where the yellow star point is. https://www.google.com/maps/d/u/0/edit?mid=17CSRYFl1B39D-lElJ2cR3x2m14ygn3CW&ll=-36.76670858962872%2C174.49151200000006&z=11 ). Samples 81-86 were all found here. I kept sample 81 which is Golddigger, sample 83 was adopted to Winnie Zeng, Courtney Armstrong and, Tyler Porteous and sample 86 was adopted to Jamie Le Roux.
We were very fortunate that in the community garden there was three very large compost bins (the bins were approximately 1.4m x 1.4m x 1.2m in length, width and height respectively) and I was able to collect 6 samples (81-86), two from each bin (one from the front and one from the back), from here which all came back with positive plaques as seen in appendix three.
As you can see in image one above, Golddigger is a siphoviridae type bacteriophage who’s tail is approximately 109.68nm long and the capsid has a diameter of approximately 48.39nm.
The full calculations for these sizes are in appendix one.
Golddigger’s morphology presents as large clear plaques which on average are 4.875mm in diameter (as seen in image two below), however the plaque sizes ranged from 3.75mm to 6mm. Golddigger and another phage (squee found by Jamie which can be seen in appendix three, plate 86) showed very similar plaque morphologies and so a gel electrophoresis was completed and is detailed below.
As you can see in image three, Golddiggers DNA isn’t easily cut by restriction enzymes. Upon analysis, the only enzymes that cut Golddigger’s DNA were ClaI, HaeIII and possibly SalI as it shows a slightly lower band (it traveled 2mm further then the uncut DNA did) then all of the other restriction enzymes.
From this analysis, it appears that Golddigger’s DNA is larger then 1kb because the uncut DNA sits 3mm higher then the 1kb marker. (3)
By completing a gel electrophoresis analysis, we were able to identify that Golddigger is distinctively different from all of the other phages found in the class. This was important because I found all of the phages used by the class, except for phage bazzle which was found by Bailey in a different location.
You can see all of the other gel electrophoresis results in appendix two.
So, despite all of the stress and tears that we had at the start of the semester with our imposter host bacteria, check out appendix four if you want to learn about that, I’m the proud mum of phage Golddigger who is beautiful and unique in it’s own special way!
Calculations showing how Golddiggers tail length and capsid diameter were worked out. Calculations are based on the calculations in reference two.
Tail length = 109.68nm
(scaled size bar / Measured size bar) = (unknown scaled size / measured size)
(50nm / 31mm) = (unknown scaled size / 68mm)
unknown scaled size = (50nm x 68mm) / 31mm
= 109.68 nm
Capsid diameter = 48.39 nm
*same calculation as above except the measured size is 30mm, therefore,
Unknown scaled size = (50nm x 30mm) / 31mm
= 48.39 nm
All six gel electrophoresis images of all of the phages found in this years class. As you can see, Golddigger shows a distinctly different banding pattern to all of the other phages found.
Plates showing the positive plaques of samples 81-86.
These two plates below were made by our laboratory technician, Jarred, who suspected that the host that we were using for the first half of the semester had become contaminated somehow. He spot tested our phages and a control phage (FancyPants, which is in green in the bottom right hand section) on the old host (left) and the a new sample of the correct host (right). As you can see, FancyPants didn’t infect the old host and so from this he concluded that the reason why we were not finding any phages is because the old host that we were using had become contaminated by something else at some point.
- Map showing where samples were taken from during phage hunt. The Yellow star is where Golddigger was found.
- Calculations based on the calculations detailed in the 2019 Course and assessment guide, 2019 by Dr Heather Hendrickson.
Protocol 8.1a, page 68.
- Gel electrophoresis set up and analysed using the phage hunt course and assessment guide 2019 by Dr Heather Hendrickson.
Protocol 10, pages 77-87.