Localization of fw2.2 mRNA expression in soybean root [abstract]
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Soybean and Rhizobia soil bacteria have a symbiotic relationship that results in nitrogen fixation, the process in which nitrogen is converted from N2 to ammonia (NH3). The bacteria induce the expression of genes responsible for the organogenesis of root nodules, the organ in which fixation occurs. Nitrogen fixation only occurs in a nitrogen-poor soil environment. The fw2.2 soybean gene was found by microarray and qRT-PCR to be upregulated during nodule development. The function of soybean fw2.2 is unknown. In tomato, the fw2.2 gene was found to negatively regulate cell division, resulting in fruit size changes of up to 30 percent. In the current studies, in situ hybridization was used to develop a time course of spatial expression of fw2.2 in soybean root. The technique used a digoxegenin (DIG)-labeled RNA probe recognized by a DIG-specific antibody coupled to alkaline phosphatase. Colorometric detection of alkaline phosphatase activity was then used to visualize the cellular location of fw2.2 mRNA. Both antisense RNA (T3 polymerase) and sense RNA (T7 polymerase) probes were used to differentiate between specific and non-specific staining. Further work will be done to determine the expression of soybean fw2.2 in roots over time as well as in seeds and root tips. We are currently optimizing the in situ hybridization technique to better illustrate the dynamics of fw2.2 expression in soybean and to further understand the biological function of fw2.2.