The last major objective our mentors set for Ruth and me this summer was conducting a bioassay. Monday, Tuesday and Wednesday we carried out the assay and analyzed the results. From past research in this lab and our work this summer, we had three pure compounds with known molecular weights and structures. With this information, we carried out serial dilutions to make seven different molar concentrations of the compound in buffer.
For hours on Monday, we measured countless wheat coeleoptiles (the primary shoot on a wheat seed made of undifferentiated cells) and placed five in each tube for each dilution... It took a while.
The coolest part was the room we worked in. The coeleoptiles need to be kept in the dark because all growth and inhibition should be due to the purified compounds in buffer. However, since it is impossible to measure and count in the dark, the room you work in has only green light. That way the plants can not absorb any light, so it is like dark for the plants. When you leave the room everything looks pink and red because your eyes need to adjust. It was a pretty exciting part of the day.
After setting up the bioassay, the coeleoptiles are incubated for 24 hours. Tuesday, we photographed each seed ... there were hundreds, and Wednesday we measured the growth using a computer program. It was exciting to see the coeleoptiles in high concentration of certain molecules were significantly shorter than those in the control or small concentrations of compound. Basically, this means we isolated bioactive compounds that inhibit growth! (It's what we hoped to accomplish this summer.)
Making many assumptions with this data, it is possible these chemicals are produced by the plants to keep other plants from growing near them and using resources (light, soil, nutrients). Of course, this will take years to prove. But showing bioactivity is a good first step! Here is a picture of the incubated wheat coeleoptiles after 24 hours of growth. Some are visibly shorter than others due to inhibition caused by the quinone compounds.
