|Fajardo, Diego -|
|Jayanty, Sastry -|
Submitted to: Journal of Visualized Experiments
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 13, 2013
Publication Date: June 7, 2013
Citation: Fajardo, D., Jayanty, S., Jansky, S.H. 2013. Rapid high throughput amylose determination in freeze dried potato tuber samples. Journal of Visualized Experiments. 80:e50407. Available: http://www.jove.com/video/50407/. Interpretive Summary: Potato starch is composed of approximately 75% amylopectin and 25% amylose. We are interested in breeding for higher amylose content, which would increase the fiber content of potato and decrease glycemic index. In order to make progress in a breeding program, we need a high throughput assay for amylose. This will allow us to screen large populations for individuals with high amylose in tuber starch. This paper describes a method using small samples in 96-well plates.
Technical Abstract: Approximately 80% of the fresh weight of a potato tuber is water; nearly all of the remaining dry matter is starch. Most of the starch (70%) is composed of amylopectin, while the remainder is amylose. The ratio between amylose and amylopectin is the most important property influencing the physical properties of starch. Amylose is a linear alpha 1-4 glucose chain, while amylopectin is a linear alpha 1-4 chain with alpha 1-6 branches. This protocol describes a colorimetric method that relies on the formation of a complex between iodine and chains of glucose molecules in starch. Iodine complexes with both amylose and long chains within amylopectin. After the addition of iodine to a starch sample, the maximum absorption of amylose and amylopectin occurs at 620 and 550 nm, respectively. The amylose/amylopectin ratio can be estimated from the ratio of the 620 and 550nm absorbance values and comparing them to a (standard curve)? in which specific known concentrations are plotted against absorption values. This high throughput, inexpensive method is reliable and reproducible, allowing the evaluation of large populations of potato clones.