Application of body time detection by molecular timetable method for quality control of postharvest fruit and vegetables

2021 Fiscal Year Research-status Report

• Project/Area Number: 21K05847
• Research Institution: Gifu University
• Principal Investigator: タンマウォン マナスィカン 岐阜大学, 応用生物科学部, 准教授 (90763673)
• Project Period (FY): 2021-04-01 – 2024-03-31
• Keywords: Circadian rhythm / Fresh produce / Quality

Outline of Annual Research Achievements

Circadian clock (CC) system produces a circadian rhythm (CR) of about 24 h a day. CC system regulates biological processes including stress responses and changes in CR state of plants. However, there is little information of this system in postharvest fresh produce. Hence, this research aims to understand the CR state of fresh produce by analysis of clock genes expression.
Soybean sprouts cultivated under dark condition were used for experiment. After harvesting, samples were collected every 2 h for 24 h. The expression levels of 47 target genes were measured by qPCR method. The time-course expression level of target clock genes was fitted to the cosine curve with a 24-h cycle. The 37 genes which expressed periodically were selected and these genes were assumed to be expressed under normal CR. The peak of the cosine curve was defined as peak time (PT). Molecular timetable (MTM) was obtained by plotting the PT and the normalized expression level of each selected gene. This plotting was then applied to the cosine curve, and the peak of the curve at this time was defined as body time (BT). Additionally, the coefficient of determination of fitting the predicted BT dataset (n=1) with the experimented BT (n=4) showed a value of 0.93, and the RMSE was 2.0 (h), so that the BT could be sufficiently estimated. Overall, it revealed that there is probability to construct a MTM by expression data of 37 time-indicating genes. Construction of MTM can be possibly conducted without relying on comprehensive gene analysis by RNA sequencing.

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

In the fiscal year 2021, from the results, there is probability to construct a molecular timetable by expression data of 37 types of time-indicating genes for soybean sprouts after harvesting. Construction of MTM can be possibly conducted without relying on comprehensive gene analysis by RNA sequencing.
In addition, sample preparation, gene expression analysis method including design of primers have been established and analysis of clock gene expression can be performed smoothly. Therefore, it has become possible to acquire the environment for conducting research and to obtain experimental data as mentioned in the plan.
Furthermore, the results of exploring expression of clock gene in soybean sprouts were analyzed and discussed and is preparing for submitting to the scientific journal.

Strategy for Future Research Activity

To investigate the effect of harvesting time on internal time, an analysis of gene expression of soybean sprouts harvested at different times during the day will be conducted. Prediction of internal time of single time sampling method by mathematical model will be examined. Influence of storage condition on clock gene expression will be also investigated and discussed.