Identification of potential molecular markers for detection of lengthy chilled storage of Prunus persica L. fruit

Abstract

Low temperature is the main strategy to preserve fruit quality post-harvest, in the supply chain. Low temperatures reduce the respiration, ethylene emission, and enzymatic activities associated with senescence. Unfortunately, peaches are sensitive to low temperatures if exposed for long periods, resulting in physiological disorders that can compromise commercial quality. Maximum damage occurs at 5 °C while at 1 °C damage is reduced. Therefore, rapid early detection methods for the distribution chain to monitor length and temperature of fruit storage are needed. The aim of this work was to identify candidate genes to develop an antibody-based marker system in peach fruit to monitor chilled storage. Two cultivars were tested: 'Sagittaria', an early ripening peach, and 'Big Top' a mid-season ripening nectarine, with delayed softening and resistance to supply-chain conditions. Both cultivars were subjected to 1 or 5 °C chilled storage for different times to simulate typical supply-chain conditions. Identification and expression of potential marker genes was assessed using a previous transcriptomic study following storage at 1 °C. Fifteen candidate genes were selected, however only seven proteins encoded were suitable as protein markers as they lack a transmembrane domain. Real-time qPCR using fruit from the subsequent year to the transcriptome was used to assess expression at both 1 and 5 °C chilled storage of five candidate genes. Four genes and the related proteins were identified that would be suitable for the development of molecular markers: a Pathogenesis-Related Bet v I family protein, a dehydrin, a Glycosyl hydrase family 18 protein and a Late Embryogenesis abundant protein.

Keywords: Cold storage; ELISA test; Gene expression; Molecular markers; Prunus persica L.

Graphical abstract

Fig. 1

Transmembrane topology prediction of proteins using DeepTMHMM tool. The x-axis represents the length of the protein and the y-axis the probability of a transmembrane domain. (A) Periplasmic beta-glucosidase, (B, C, D) Thaumatin family, (E, F) Glycosyl hydrolases, (G) fatty acid desaturase, (H) Acid phosphatase.

Fig. 2

Gene expression changes in Sagittaria peach and Big Top nectarine during cold storage treatment at 1 °C at 0, 1, 5, 7 and 14 d followed by 36 h recovery at ambient temperature (22 °C) in 2017 based on transcriptomic analysis. Differentially expressed genes (DEGs) during the time course are expressed as read counts and obtained by ImpulseDE2. (A) Glutamine synthetase beta-Grasp domain, (B) Glycosyl hydrolases family 18, (C) Late Embryogenesis Abundant (plants) LEA-related, (D) Pathogenesis related protein Bet v I family, (E) Dehydrin. Different letters indicate significant differences among cultivars considering all time points. Statistical analyses were performed using Two-way ANOVA and Tukey's ranked test (P < 0.05). Data are the mean ± SE; n = 3.

Fig. 3

Real-time PCR analysis of selected DEGs in Sagittaria peach and Big Top nectarine during cold storage treatment at 1° (A, C, E, G) and 5 °C (B, D, F, H) at 0, 1, 5, 7, and 14 d followed by 36 h recovery at ambient temperature (22 °C). Relative expression (2^−ΔCt): (A) and (B) Glycosyl hydrolases family 18; (C) and (D) Late Embryogenesis Abundant (plants) LEA-related; (E) and (F) Pathogenesis related protein BET v I family, (G) and (H) Dehydrin. Different letters indicate significant differences among cultivars considering all time points. Statistical analyses were performed using Two-way ANOVA and Tukey's ranked test (P < 0.05). Data are the mean ± SE; n = 3.