Monitoring Apricot (Prunus armeniaca L.) Ripening Progression through Candidate Gene Expression Analysis

Abstract

This study aimed at the monitoring of the apricot (Prunus armeniaca L.) ripening progression through the expression analysis of 25 genes related to fruit quality traits in nine cultivars with great differences in fruit color and ripening date. The level of pigment compounds, such as anthocyanins and carotenoids, is a key factor in food taste, and is responsible for the reddish blush color or orange skin and flesh color in apricot fruit, which are desirable quality traits in apricot breeding programs. The construction of multiple linear regression models to predict anthocyanins and carotenoids content from gene expression allows us to evaluate which genes have the strongest influence over fruit color, as these candidate genes are key during biosynthetic pathways or gene expression regulation, and are responsible for the final fruit phenotype. We propose the gene CHS as the main predictor for anthocyanins content, CCD4 and ZDS for carotenoids content, and LOX2 and MADS-box for the beginning and end of the ripening process in apricot fruit. All these genes could be applied as RNA markers to monitoring the ripening stage and estimate the anthocyanins and carotenoids content in apricot fruit during the ripening process.

Keywords: Prunus armeniaca L.; breeding; fruit quality; metabolomics; multiple linear regression; predictive models; ripening; transcriptomics.

Figure 1

Phenological and fruit quality traits boxplots (fruit weight, skin color, flesh color, soluble solids content, acidity (pH), firmness, chlorophyll a, chlorophyll b, anthocyanins, carotenoids and ripening date) evaluation of the nine cultivars assayed, ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at physiological ripening. Boxplots are filled with the fruit color of each cultivar.

Figure 2

Principal Component Analysis (PCA). From left to right: representation of the variable contribution to dimension 1 and 2, variable contribution and dimension correlation with the first five dimensions and percentage of explained variance for the first ten dimensions performed over phenological and quality traits (fruit weight, skin color, flesh color, soluble solids content (SSC), acidity (pH), firmness, chlorophyll a, chlorophyll b, anthocyanins, carotenoids and ripening date) evaluated in apricot fruit at physiological ripening. The cultivars evaluated were ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z108-38′.

Figure 3

Principal Component Analysis (PCA) performed over the phenological and fruit quality traits (fruit weight, skin color, flesh color, soluble solids content (SSC), acidity (pH), anthocyanins and carotenoids content and ripening date (RD)) of nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at physiological ripening. Symbols represent each cultivar and are colored as the real fruit color previously described. This color legend will be used for all bar plots in this study.

Figure 4

Anthocyanins and carotenoids content bar plots of the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at three different ripening stages based on their skin ground color and firmness: green fruit (Sample A), during color change (Sample B) and at physiological ripening (Sample C).

Figure 5

Expression dynamics barplots of the genes involved in anthocyanin biosynthesis (4CL, ANS, bHLH, C4H, CAD1, CHS, DFR, F3′5′H, MYB10, UFGT and WD40) in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at three different ripening stages based on their skin ground color and firmness: green fruit (_A), during color change (_B) and at physiological ripening (_C).

Figure 6

Expression dynamics barplots of the genes involved in carotenoids biosynthesis and degradation (CCD1, CCD4, CrtL, DXS and ZDS) in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at three different ripening stages based on their skin ground color and firmness: green fruit (_A), during color change (_B) and at physiological ripening (_C).

Figure 7

Expression dynamics barplots of the genes described as metabolites transporters GST and MATE, in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at three different ripening stages based on their skin ground color and firmness: green fruit (_A), during color change (_B) and at physiological ripening (_C).

Figure 8

Barplots of the expression dynamics of others genes involved in the regulation of ripening progress, as bZIP, EDR1, LOX2, MADS-box, NAC, psbP1 and WRKY, in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z 108-38′ at three different ripening stages based on their skin ground color and firmness: green fruit (_A), during color change (_B) and at physiological ripening (_C).

Figure 9

Pearson correlation plot for the relative gene expression evaluated by using RT-qPCR of genes involved in anthocyanins and carotenoids biosynthesis and ripening process progression in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z108-38′ at three ripening stages green fruit, during color change and at physiological ripening. The assayed genes included DFR, LOX2, GST, CCD4, DXS, bHLH, MYB10, bZIP, MADS-box, CAD1, CrtL, ZDS, ANS, WD40, C4H, CHS, MATE, UFGT, CCD1, WRKY, F3′5′H, CL4, psbP1, NAC and EDR1.

Figure 10

Pearson correlation (r coefficient) network for relative expression by RT-qPCR of genes involved in anthocyanins and carotenoids biosynthesis with correlation values over 0.7. The plotted genes were C4H, CAD1, CHS, MYB10, CrtL, CCD1, UFGT, WD40, WRKY, ZDS, bZIP, DXS and MADS-box.

Figure 11

Principal Component Analysis (PCA) of relative expression from anthocyanins content MLR model. From left to right: representation of the variable contribution to dimensions 1 (Dim1) and 2 (Dim 2), variable contribution and dimension correlation with the first five dimensions and percentage of explained variance of the first ten dimensions performed over the relative expression of the genes involved in anthocyanins biosynthesis (ANS, bHLH, bZIP, C4H, CAD1, CHS, CL4, DFR, EDR1, F3′5′H, GST, MATE, MADS-box, MYB10, NAC, LOX2, UFGT, WD40 and WRKY) evaluated in apricot fruit during the ripening progression in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z108-38′ at three ripening stages green fruit, during color change and at physiological ripening.

Figure 12

Principal Component Analysis (PCA) of relative expression from carotenoids content MLR model. From left to right: representation of the variable contribution to dimensions 1 (Dim1) and 2 (Dim2), variable contribution and dimension correlation with the first five dimensions and percentage of explained variance of the first ten dimensions performed over the relative expression of the genes involved in carotenoid biosynthesis (bZIP, CCD1, CCD4, CrtL, DXS, GST, MATE, MADS-box, NAC, LOX2 and ZDS) evaluated in apricot fruit during the ripening progression in the nine cultivars ‘Bergeron’, ‘Cebasred’, ‘Currot’, ‘Dorada’, ‘Estrella’, ‘Goldrich’, ‘Moniquí’, ‘Valorange’ and ‘Z108-38′ at three ripening stages green fruit, during color change and at physiological ripening.

Figure 13

Ripening stages before stone hardening based on their skin ground color and firmness assayed in this study in the light yellow cultivar ’Moniquí’ and the orange cultivar ‘Goldrich’: green fruit (Stage A), during color change (Stage B) and at physiological ripening (Stage C).