BO5 - Developing and utilizing qPCR techniques to study the relationships between polyphenol oxidase and polygalacturonase and oxidation in Granny Smith apples
SCURS Disciplines
Biology
Document Type
General Presentation (Oral)
Invited Presentation Choice
Not Applicable
Abstract
Polymerase chain reaction (PCR) is a technique that can amplify copies of nucleic acids sequences quickly. qPCR (quantitative PCR) is a type of PCR that actively measures the amount of DNA or RNA . The ability of qPCR to quantify results allows for a wider range of experiments to be completed. Since RNA, when compared to DNA, is more prone to degeneration and instability following isolation from tissues, it is appropriately suited for qPCR analysis. Previously, no established protocol for qPCR had been developed for use in our undergraduate curriculum, despite owning a Chai Open qPCR system. First, qPCR calibration was established using known values of diluted DNA concentrations to compare results obtained from qPCR. Then, as proof of concept, three strawberry genes associated with fruit ripening, ERF3, ERF61, ERK71a, were quantified to determine the reproducibility of the qPCR system. Previous research in the labratory suggested activity of the antioxidant and browning enzyme polyphenol oxidase (PPO) varied among species of apples. However, levels of PPO expression were never confirmed. An experimental question was proposed to use PPO and polygalacturonase (PG) to determine the degree of browning in apples over a set period of time, as both genes are active in apple oxidation. Granny Smith apples were cut and set out for 0 hours, 1 hour, 6 hours, and 24 hours at room temperature. Using established techniques from previous strawberry experiments, mRNA was isolated from apples and qPCR performed to quantify PPO and PG. These genes were established, and the genes were found to increase in expression over time. A total fold change of four was observed at the 24-hour sample when compared to the 0-hour sample. Data demonstrates levels of PPO expression that correlate with PPO activity. Also, the establishment of qPCR techniques at our university allows the qPCR machine to be utilized in areas of the biology curriculum such as cell biology, microbiology, or molecular biology, and for independent research. The understanding of the relationship between PPO gene expression, enzymatic activity, and browning can provide for future research uncovering the pathways behind browning and oxidation in apples.
Keywords
qpcr, polyphenol oxidase, polygalacturonase, Granny Smith, oxidation, RNA
Start Date
10-4-2026 3:25 PM
Location
CASB 101
End Date
10-4-2026 3:40 PM
BO5 - Developing and utilizing qPCR techniques to study the relationships between polyphenol oxidase and polygalacturonase and oxidation in Granny Smith apples
CASB 101
Polymerase chain reaction (PCR) is a technique that can amplify copies of nucleic acids sequences quickly. qPCR (quantitative PCR) is a type of PCR that actively measures the amount of DNA or RNA . The ability of qPCR to quantify results allows for a wider range of experiments to be completed. Since RNA, when compared to DNA, is more prone to degeneration and instability following isolation from tissues, it is appropriately suited for qPCR analysis. Previously, no established protocol for qPCR had been developed for use in our undergraduate curriculum, despite owning a Chai Open qPCR system. First, qPCR calibration was established using known values of diluted DNA concentrations to compare results obtained from qPCR. Then, as proof of concept, three strawberry genes associated with fruit ripening, ERF3, ERF61, ERK71a, were quantified to determine the reproducibility of the qPCR system. Previous research in the labratory suggested activity of the antioxidant and browning enzyme polyphenol oxidase (PPO) varied among species of apples. However, levels of PPO expression were never confirmed. An experimental question was proposed to use PPO and polygalacturonase (PG) to determine the degree of browning in apples over a set period of time, as both genes are active in apple oxidation. Granny Smith apples were cut and set out for 0 hours, 1 hour, 6 hours, and 24 hours at room temperature. Using established techniques from previous strawberry experiments, mRNA was isolated from apples and qPCR performed to quantify PPO and PG. These genes were established, and the genes were found to increase in expression over time. A total fold change of four was observed at the 24-hour sample when compared to the 0-hour sample. Data demonstrates levels of PPO expression that correlate with PPO activity. Also, the establishment of qPCR techniques at our university allows the qPCR machine to be utilized in areas of the biology curriculum such as cell biology, microbiology, or molecular biology, and for independent research. The understanding of the relationship between PPO gene expression, enzymatic activity, and browning can provide for future research uncovering the pathways behind browning and oxidation in apples.