Abstract:【Objective】Actinidia eriantha is a unique kiwifruit species of Actinidia spp. The fruit of A. eriantha is rich in nutrients and ascorbic acid (vitamin C). However, the flavor quality of most wild A. eriantha resources is lower than that of the cultivars from A. chinensis or A. deliciosa mainly due to the low accumulations of soluble sugars. Fortunately, a high sugar type kiwifruit cultivar, A. eriantha Ganlv 1, has been developed by our research group recently. In order to explore the formational mechanism of the high accumulation of soluble sugars, and to identify the related key metabolic enzyme and key gene in fruit of A. eriantha Ganlü 1 at the ripening stage, A. eriantha Ganlv 1 and another low sugar type kiwifruit cultivar, A. eriantha Ganlv 6, were selected as the research materials in this study. These two cultivars possess similar genetic background and they both originated from the wild A. eriantha resources of Magu Mountain in Nancheng County, Jiangxi Province.【Methods】The grafted vines of A. eriantha Ganlü 1 and A. eriantha Ganlü 6 were planted in an orchard of the Fengxin County Bureau of Agriculture and Rural Affairs, which located in Fengxin County, Jiangxi Province. In 2021, at physiological maturity stage when the soluble solids content of fruits from each cultivar reached more than 7.0%, fruits without pests, diseases and mechanical damage were collected randomly from selected vines. After collection, the fruits were placed in an ice box and transported to the laboratory for further treatment. For each cultivar, three uniform vines were used as three biological replicates. Twenty fruits were collected from each vine. For fresh fruits, fruit weight was measured using an electronic balance; fruit longitudinal diameter and fruit transverse diameter were measured respectively with a vernier caliper. Fruit shape index was obtained using the ratio of the longitudinal diameter to the transverse diameter. Then those fruits were stored at room temperature until soft-ripe stage. For those fresh fruits, soluble solids content (SSC) was measured with a portable refractometer, and dry matter content (DM) was measured using fruit slices. For the remaining fruits, after removing the peel, seeds and core, the flesh was separated, sliced, frozen with liquid nitrogen, and stored in a refrigerator at –80℃. For the frozen flesh samples, the soluble sugar content (SS) was determined using anthrone sulfuric acid colorimetry; the titratable acidity (TA) was determined by titration with NaOH; and the ascorbic acid (AsA) content was determined by the 2,6- dichlorphenolindophenol titration method. Sugar-acid ratio was calculated by the ratio of the SS to the TA. Also, glucose (Glu), fructose (Fru), and sucrose (Suc) were extracted from the frozen samples in 80% ethanol, and their contents were determined by high-performance liquid chromatography. Total soluble sugar content (TS) was obtained by calculating the sum of Glu, Fru, and Suc. Sweetness value (SV) was calculated based on the contents of Glu, Fru, Suc, as well as their corresponding coefficients. Besides, for the frozen samples, activity assays of the main enzymes involved in Suc metabolism, including sucrose phosphate synthase (SPS), sucrose synthase (SUSY), acid invertase (AINV), and neutral invertase (NINV), were carried out using their corresponding kits. Furthermore, the relative expression levels of the related genes encoding those enzymes, including AeSPS, AeSUSY, AeAINV, and AeNINV, were analyzed by qRT-PCR method for A. eriantha Ganlü 1 and A. eriantha Ganlü 6. In addition, correlation relationships among different indicators were analyzed using bivariable analysis following the Pearson method. Those indicators included SS, SSC, SV, TS, Glu, Fru, Suc, the activities of SPS, SUSY, AINV, NINV, and the relative expression levels of AeSPS, AeSUSY, AeAINV, and AeNINV.【Results】Fruit size, fruit shape index, and fruit weight were significantly higher in A. eriantha Ganlü 1 than that in A. eriantha Ganlü 6. Most of the indicators related to internal fruit quality were also significantly higher in A. eriantha Ganlü 1 than that in A. eriantha Ganlü 6. Those indicators included SSC, SS, sugar-acid ratio, and AsA content. Besides, higher contents of Fru, Suc, TS, and higher SV were detected in fruits of A. eriantha Ganlü 1 at the ripening stage. Additionally, in fruit of Ganlü 1, the activities of SPS and SUSY were both significantly higher than that of Ganlü 6; however, the activities of AINV and NINV were significantly lower in fruit of Ganlü 1. Meanwhile, the relative expression levels of AeSPS and AeNINV were significantly higher in fruit of Ganlü 1 than that of Ganlü 6. Moreover, soluble sugar content was positively correlated to SSC, SV, the contents of TS, Fru, Suc, and the activities of SPS and SUSY; while it was negatively correlated to Glu content, and the activities of AINV and NINV. Furthermore, the relative expression level of AeSPS was found to be positively correlated to both Suc content and SPS activity.【Conclusion】The difference in sucrose accumulation level is the dominant factor that results in the great difference in soluble sugar content between the cultivars of A. eriantha Ganlü 1 and A. eriantha Ganlü 6. And sucrose phosphate synthase is the key metabolic enzyme that positively regulates sucrose accumulation in ripe fruit of A. eriantha culitvars at the ripening stage. Besides, we speculated that AeSPS is the key gene which contribute to the biosynthesis of sucrose by increasing SPS activity in fruits of A. eriantha Ganlü 1 at the ripening stage.
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