Biomass of different parts
The biomass of post-grafting generation of G1, G2, and G3 increased in the root, stem, and leaf biomass. The order of biomass from highest to lowest is as follows: G3 > G2 > G1 > CK. Different numbers of grafting resulted in no significant difference in root biomass of grafted progeny compared to control (P>0.05; Fig. 1a). Moreover, G1, G2, and G3 treatment increased shoot biomass by 21.10%, 27.58%, and 34.07% (p < 0.05; Fig. 1b), compared to ungrafted progeny.
Photosynthetic pigment content
The chlorophyll (chlorophyll a and chlorophyll b) content in the leaves increased with the number of grafting compared to the control, with an increase in chlorophyll content of 0.84%, 9.05%, and 17.46% in the grafted progeny G1, G2 and G3, respectively (Fig. 2a), and an increase in the content of carotenoids of 57.78%, 65.33% and 86.67%, respectively (Fig. 2b). However, the chlorophyll content of G1 did not change significantly (P > 0.05).
Antioxidant enzyme activity and soluble protein content
Except for SOD activity, CAT, POD activity, and soluble protein content were increased in leaves of different generations compared with the control. The G1 and G2 treatments had no significant effect on CAT activity (P > 0.05), whereas G3 increased CAT activity by 55.56% (P < 0.05; Fig. 3c). Compared with CK plants, G1, G2, and G3 treatments increased POD activity by 31.80%, 33.21% and 87.49% (P < 0.05; Fig. 3a) and soluble protein content by 10.15%, 16.92% and 23.16% (P < 0.05; Fig. 3d), respectively. SOD activity decreased in different grafting treatments, with lower activity in G3, followed by G1 and G2 (p < 0.05; Fig. 3b).
Se content and transport in post-grafting generations
The selenium content in the roots increased progressively with the number of grafts. The selenium content of seedlings G1, G2, and G3 significantly increased by 66.43%, 45.20%, and 32.21%, respectively (P < 0.05; Fig. 4a) compared with the ungrafted treatment. The highest selenium content of 28.06 mg kg− 1 was found in G3. However, a decreasing trend was observed in the selenium content of stems, although the decrease in selenium content was not obvious in seedlings G1, which was significantly reduced by 5.23% and 8.67% for G2 and G3, respectively, at 7.43 mg kg− 1 and 7.16 mg kg− 1 (Fig. 4b). In addition, the selenium content in leaves decreased gradually with increasing number of grafts. selenium content in leaves of G2 and G3 decreased significantly by 11.22% and 17.50% to 13.29 mg kg− 1 and 12.35 mg kg− 1, respectively, compared with the control (Fig. 4c). The translocation factor (TF) decreased significantly with increasing number of grafts. G1, G2, and G3 were significantly reduced by 27.64%, 39.50%, and 48.83% (P < 0.05; Fig. 4d) compared to the control.
Se accumulation content in post-grafting generations
Continuous grafting increased the accumulation of selenium in the root, with significant increases of 33.59%, 50.00%, and 74.48% in G1, G2, and G3 treatments, respectively, compared to the control (P<0.05; Figs. 5a). After grafting, the selenium accumulation in the stem also increased, but the accumulation of selenium did not increase significantly (Fig. 5b).
Correlation analysis
Root Se content, leaf Se content, shoot Se accumulation, and root Se accumulation were positively correlated with root biomass, stem biomass, leaf biomass, chlorophyll content, carotenoid content, POD activity, CAT activity, and soluble protein content, whereas they were negatively correlated with SOD activity and TF (Fig. 6). In addition, leaf Se content was not correlated with chlorophyll content (relative coefficient = 0). Stem Se content was negatively correlated with root biomass, stem biomass, leaf biomass, chlorophyll content, carotenoid content, POD activity, CAT activity, and soluble protein content, while it was positively correlated with SOD activity and TF. Positive correlations were also found between root Se content, leaf Se content, aboveground Se accumulation, and root Se accumulation. On the contrary, stem Se content was negatively correlated with aboveground Se accumulation and root Se accumulation.
Principal Component Analysis (PCA)
Principal component analysis (PCA) was performed on seedlings progeny of different grafting treatments (CK, G1, G2, G3) based on the measured indicators (Fig. 7). Two principal components were selected based on eigenvalues (eigenvalues greater than 1). The cumulative contribution of the first and second principal components was 90.2%, indicating that the first two principal components could reflect most of the information contained in the original indicators. The PCA scores reflect the effects of continuous grafting on tomato progeny biomass, chlorophyll content, antioxidant enzyme content, selenium accumulation, and selenium transportation factor (TF) under 10 mg kg− 1 selenium treatment. Triple grafting treatments contained in the upper right quadrant of PC1 resulted in progeny with high root biomass, chlorophyll II content, CAT, POD activity, soluble protein content, and high root Se accumulation. The double grafting treatment (G2) was located in the lower right quadrant of PC1 and had higher leaf biomass, shoot biomass, carotenoid content, and root Se content after grafting. Finally, the lower and upper left quadrants outline the ungrafted plants (CK) and single grafted (G1) treatments, characterized by the lowest stem and leaf se content but higher SOD activity.
Grey relational analyses
From the gray correlation diagram, it can be seen that there were correlations between shoot Se accumulation and several factors such as biomass, plant Se content, root Se accumulation, photosynthetic pigment content, antioxidant enzyme activity, and soluble protein content (Fig. 8). The gray correlation coefficients of carotenoid content and SOD activity were lower than 0.60, while the correlation coefficients of other indexes with shoot Se accumulation were higher than 0.6. Thus, it can be seen that the biomass, plant Se content, root Se accumulation, and shoot Se accumulation were closely related to each other.