An experiment to determine if the DCl4 or RDR6 genes control the gravitropic response of Arabidopsis Thaliana Aiden Rohde, Michael Savory, Lindsey Yost, Hunter Willis Abstract The PI’s aim to determine whether or not the RNA silencing genes, DCL4 or RDR6, have a impact on the responses of Arabidopsis Thaliana to a shift in orientation. Beyond assisting in the attempt to find, by process of elimination, the function of the above two genes, this research will provide insight into the genetic processes required to grow plants in variable gravity, such as they might be subjected to in space. Said research will by instrumental in practices which are designed to increase the health of crop plants such as the growth of tomatoes in a inverted position. Procedure: The PI’s planted a total of 12 pots, each containing four separate plants. Half were treatment and half were control, and of each group two pots were DLC4, two RDR6, and two Wild-type. The plants were watered Mondays and Thursdays and one teaspoon of fertilizer was added per gallon of water. Three weeks after planting the Arabidopsis, they had grown stalks of sufficient height to perform the gravitropic treatment. The first test was intended to analyze the short term response to a gravitropic treatment. The experiment was performed by setting the plants being treated on their sides so that the stalks were parallel to the ground. Then a cardboard box was placed over the plants to eliminate light as a variable so that the only variable being tested was the effect of gravity. The control plants were also placed underneath a cardboard box to simulate similar environmental factors with the exception of the gravitropic treatment. Every 5 minutes for 35 minutes the angle of the stalks were measured to analyze the extent of gravitropic response in the allotted time. The second test was designed to analyze long term response to gravitropic treatment. The same procedure was followed as done for the first test with the exception being that intervals between measurements were increased to 1 hour. Data and Analysis The PI’s determined that in both the long and short rounds of gravitropic trial the DCL4 mutants tended to respond with more angulations in the same time frame. However, both the RDR6 and Wild-Type plants displayed more total change in orientation than DCL4. RDR6 and Wild-Type plants had reasonably similar levels of change in both trials. From these results the PI’s determined that DCL4 had an effect on the gravitropic response of Arabidopsis in that it increased its rate. Because RDR6 behaved similarly it has been determined by the Pi’s that a mutation in the RDR6 gene will have no effect on the gravitrophic response of Arabidopsis. Background Gravitropism- The growth of a plant in response to gravity. As initially discovered by Charles Darwin, plant roots show ‘positive gravitropism’ in which they grow in the direction of gravitational pull and plant stems show ‘negative gravitropism in which they grew opposite the pull of gravity. RNA Interference- (RNAi) is the silencing of gene expression, triggered by the presence of double- stranded RNA homologous to portions of the gene. This mode of gene silencing occurs through the degrading of a target gene’s mRNA or by the blocking of translation on an active mRNA. Illustrated in Figure 2 is the process of RNA interference occurs. First a RDR, RNA Dependent RNA polymerase, makes a two stranded double helix of RNA. In the present research the PIs are using the RDDR6 gene. Then the DCL4, the last in a group of four dicer enzymes, recognizes and dices double- stranded RNA into small interfering RNAs (siRNAs) about base pairs long. The siRNAs bind to proteins which assemble the RNA-induced silencing complex (RISC). Energy from ATP allows the unzipping and therefore activation of the RISC. After activation the RISC can recognize and bind to the target mRNA and cleave the mRNA strand leaving it unable to produce the designated protein. From: research.com/content/6 /2/78/figure/F1?highres =y Figure 2 From: Figure 2 From: