On Monday (June 30, 2014) the OUTPACE began their lab session by inspecting our bacterial and fungal isolations from last Friday. The fungal growth from our samples is progressing as expected and will be further examined on Wednesday. For today’s lab our bacterial isolations must be processed further by collecting single colonies and placing them into a new agar plate by utilizing the quadrant streak method.
Students were also required to bring their own infected plant materials into the lab in order to be tested for certain pathogens. Students were asked to give a prediction of about the type of pathogen that they could have and tasked with isolating the pathogen. My partner, Maggie brought in a diseased potato, which we both used for today’s exercise.
Other samples included a mango, a peach, raspberries and a variety of leaves from our backyards.
The students performed the bacterial and/or fungal isolation method (depending on their material) performed last Friday in which four squares of infected tissue were cut out and sterilized in a 10% Clorox solution. The samples were transferred to a small tube containing water and a grinding bead to be grounded into a liquefied solution. Here is us grinding the samples on a grinding robot:
The solution was diluted three times in 1:10, 1:100, and 1:1000 solutions and placed in three specific plates and spread across the agar with the help of small beads. We concluded today’s lab with a small interview with each OUTPACER and their own personal assessment of the program overall.
On Tuesday (July 1, 2014) there was a lecture on phytoparasitic nematodes and their impact on plant species. We learned that a small percentage are parasitic in nature and are biotrophic. Nematodes are able to produce 300-400 eggs and over the period of one season can produce over 8 billion offspring, which is why they are a substantial problem, especially in agriculture. Nematodes are usually found in soil and will make their way to a root in order to absorb nutrients. The major points of the lecture were that they can live inside (endo-parasitic) or outside (endo-parasitic) and use a stylet in order for them to burrow within the root of the host and cause major problems such as premature wilting, stunted growth, and galling or cysting of roots. Some plant defenses have been observed, but due to natural selective factors there is no real distinct prevention of these parasites. More scientific research is needed to address the genetic basis of plant nematode resistance.
On Wednesday (July 2, 2014) the OUTPACE students were back in the lab continuing from our previous work on Monday. The session began with a brief explanation by Dr. Mukhtar explaining the importance of long term preservation and how using this method allows scientist to study strains of pathogens for future use. We used our inoculated bacteria that we prepared Monday for this exercise.
Using pipettes, 1mL of 80% glycerol is directly added into the bacterial tube and mixed. A small cryogen tube was passed out for each student and 1mL of the bacterial glycerol solution was poured into the tube. Once sealed, the tubes are placed into a liquid nitrogen medium where they will instantly freeze and safely preserved. Here is a picture of the cryo-preserved bacterial glycerol stocks in liquid nitrogen.
Our next exercise involved the fungal cultures we prepared on Monday. After three days of growth, the cultures had undergone favorable progress. However, in order to further purify our samples, a small area of fungi was to be transferred into a new agar place. Sterile instruments and new V8 agar plates were passed out to the students. We hope to see our new fungal plates grow over the long weekend.
Lastly, the students were handed back their infected plate samples from home to observe and further process. We took samples from a single colony and transferred it into a new agar plate by using the quadrant streak method.
Although today’s week was shorter due to the July 4th Holiday, we are always learning new material and methods in the lab and future careers. Look forward to week 6 as we continue our work here in OUTPACE.