Looking for Phages

By Fion Shiao

On the day of convocation, my friends and I went to a walking trail near school where I decided to obtain my dirt sample. I chose a spot full of vegetation thinking I’M GOING TO GET A LOT OF PHAGES! Being extremely-self conscious, I sat there as I used a spoon to transfer dirt into my plastic bag.

On the first day of lab, I was very excited about getting my own lab coat, my own lab bench and my own glasses. Following the SEA Lab Manual’s instructions, I extracted phage from my soil sample by adding PB buffer and filtering it. I then did direct plating by infecting my phage sample with M. smegmatis and plating them on agar plates. At the same time, I also prepared enrichment culture.

The next lab day, I checked my direct plating for results, and I sadly had no phages. Fortunately, there was still hope in my enrichment culture. I diluted my enrichment culture into different concentrations and plated them. Checking my results the next class, some of my diluted plates had phages. I then picked four distinct plaques with different morphologies and streaked them on plates before adding M.smegmatis and Top Agar with the aim of isolating phages. As an extremely creative person, I of course named my plaques A, B, C and D.

I thence began my endless journey of streaking. In this ongoing process of streaking, I unfortunately had to abandon A and B. After looking at my third streaking, my type C and D plaques still had different morphologies, so from C and D, CA , CB , DA and DB were created. After streaking for four times, one of the TAs pointed out how my plaques are all too concentrated. As it turns out, having too many phages is not always a great thing. He advised me to try a method, which worked for another student with the same problem. Following the method, I used a stick to dab on the plaque I wanted to isolate and swirled the stick in 100 μL of Phage Buffer before streaking. Last class, I checked my plates, and the dilution technique worked!! However, my plaques’ size still varied so much that I streaked a big plaque and a small plaque from each plate to test whether the sizes vary because of the phage’s morphology or because the different sized plaques come from different types of phages. With a bad habit of not throwing out my old plates, my stack of plates has been growing higher each class while I await for my plates to yield plaques with identical morphologies.

Advertisements
This entry was posted in From the Phage Hunters. Bookmark the permalink.