Chris Nye and Dr. Jeff Maughan, Plant and Wildlife Sciences
Chenopodium quinoa, commonly known as quinoa, is a vital crop to the subsistence farmers of the Altiplano region of South America. Quinoa has several unique characteristics which contribute to its ability to grow in a variety of harsh conditions found in the high altitudes and heat of South America. Quinoa has high drought tolerance, salt tolerance, as well as a natural resistance to birds and other pests. This protection comes from a coating of a wax-like substance, called saponin, found on the surface of quinoa seeds (Ward, 2001).
While saponins are necessary to provide quinoa with the protection it needs from pests, it has a bitter taste which reduces palatability and can also negatively impact nutritional absorption (Chauhan et Al., 1992). The layer of saponin must first be removed before the seeds can be used in food products. Various processes have been developed to remove this layer from seeds including mechanical abrasion and soaking in water. However, these processes reduce the economic benefits that farmer’s could receive from growing quinoa. To assist in breeding programs that would result in quinoa free of bitter saponins, we focused on finding genes responsible for the biosynthesis of saponins (Kurosawa et Al., 2002).
The first step in this project was to successfully grow offspring of a bitter and sweet varieties of quinoa. This required careful observation, watching for pests, watering, and providing fertilizer for the plants. In an attempt to approximate seed age, we marked the flowers on the plant according to the day they first opened. We then proceeded to harvest the seeds at various stages of development. The method of seed harvesting we used was labor intensive, as each seed had to be individually extracted from its covering and put on dry ice. The first stage, known as “watery”, were harvested while the seeds were still very small and green. The second stage, “milky”, was harvested as the seeds became larger, and produced a milky substance upon puncture. Finally, seeds were harvested at the “pasty” stage right before they became fully mature. With these several stages of seed maturity we proceeded to extract RNA in order to assess which genes are turned on and off.
We attempted to use a RNA extraction method where we first freeze-dried our seed samples and crushed them before extracting RNA. Only after extracting RNA from most of our samples and looking for purity in a formaldehyde agarose gel did we realize that this method was highly susceptible to contamination and degradation by nuclease. We then changed our method and began to extract using the RNeasy kit from Qiagen. The quality of our RNA samples greatly improved, but we had lost most of our “watery” stage seeds to the freeze-dried method. As a result, we had to replant and wait for the quinoa to grow for us to harvest that stage once more. We also determined that our seed harvesting methods exposed the seeds to contamination and degradation by removing them from their protective layers and delaying freezing. To compensate for this the new method of harvesting is to pluck off the seeds and immediately place them in liquid nitrogen.
Sadly, I graduated before seeing the end of the project and observing the gene expression profiles by microarray. I have kept in touch with the group and have learned that they are almost ready to perform the microarray with the second crop of quinoa seeds. Hopefully the results of this work will be published before the end of the year. This research will provide basic understanding into the gene expression found in quinoa seeds and lead to future discoveries and improvements in quinoa seed quality and crop performance.
References
- Chauhan GS, Eskin NAM, Tkachuk R (1992) Nutrients and anitnutrients in quinoa seed. Cereal Chem 69:85–88
- Kurosawa Y, Takahara H, Shiraiwa M (2002) UDP-glucuronic acid:soyasapogenol glucuronosyltransferase involved in saponin biosynthesis in germinating soybean seeds Planta 215: 620–629
- Ward SM (2001) A recessive allele inhibiting saponin synthesis in two lines of Bolivian quinoa (Chenopodium quinoa Willd.). Heredity 92:83–86.