An exploratory analysis of gene expression in dwarf mistletoe (Arceuthobium americanum) using qRT-PCR

Dwarf Mistletoe (Arceuthobium americanum) exhibits some very interesting characteristics, such as being a hemi-parasitic plant-on-plant parasite. It’s most interesting and most characteristic feature is its seed dispersal via explosive discharge. This seeds dispersal method is very reminiscent of a cannon shooting a cannonball. The mechanism of how the small female fruit are able to do this is currently unknown with different theories trying to explain it. The theory with the most backing is a thermogenesis theory. As the fruit is monumentally smaller than the trees it infects it has a theoretical infinite source of water to draw from. Coupled with the fruits ability to trap water with a waxy viscin and produce heat via thermogenesis it is thought that the fruit will use water pressure to propel its seed. As water is drawn in, heat is generated, expanding the water and creating pressure as the water is trapped by the wax until it ruptures the cuticle and expels the seed. This is the basis of this study as it aims to investigate the genetic side of this theory. A previous study using the Affymetrix microarray heterologous probe approach revealed the potential presence and expression of key developmental genes. Four of these key genes for this theory were studied: 14-demethylase, Alkaline Invertase, Cer1, and Shine 3. Each of these genes was run through quantitative real-time polymerase chain reaction with primers designed upon plant various genetic libraries (tobacco, wine grapes, maize, Arabidopsis, etc.). Various cDNA sample extracted from dwarf mistletoe were also ran. These samples were extracted over the growing season to the time of seed dispersal from mature (3rd year) female fruits. RuBisCO (Ribulose-1,5-bisphosphate carboxylase/oxygenase) shown to be constitutively expressed and was used as reference gene. By comparing the Cq values over the growing season of each gene a picture of the genetic expression patterns of the genes could be seen. 14-demethylase was shown to have Cq values of June = 38.07, July = 35.69, and August = 36.28. This shows the genes expression increasing from June to July where it peaks and falls off in August. The Alkaline Invertase showed a similar pattern of expression with June = 30.32, July = 26.83, and August = 31.47. The Cer1 and Shine3 genes showed inconclusive results after several runs and many adjustments to the procedure. From the data collected, it may be possible that that the 14-demethylase and Neutral Invertase are preparative, helping with protection and energy storage prior to dispersal respectively. To solidify any speculations around these results further research into the genetics of this plant needs to be conducted.