毛花猕猴桃成熟期果实品质和蔗糖代谢特征分析

Analysis of the fruit quality and the metabolic characteristics of sucrose in fruit of Actinidia eriantha at ripening stage

[目的]探究果实成熟期可溶性糖含量差异大的原因,以挖掘毛花猕猴桃果实蔗糖代谢关键酶和关键基因。[方法]以高糖型赣绿1号和低糖型赣绿6号毛花猕猴桃品种为研究材料,测定果实生理成熟期外观指标;测定软熟期可溶性固形物含量、干物质含量、可溶性糖含量、糖组分含量、甜度值、蔗糖代谢酶活性和代谢酶基因相对表达量等指标;分析不同指标间的相关性。[结果]果实成熟期可溶性糖含量与蔗糖含量呈显著正相关;蔗糖含量与蔗糖磷酸合酶活性及其编码基因AeSPS表达量呈显著正相关。[结论]蔗糖是引起毛花猕猴桃果实可溶性糖含量差异大的主要原因,蔗糖磷酸合酶是影响果实成熟期蔗糖代谢的关键酶,推测其编码基因AeSPS是调控果实成熟期蔗糖合成的关键基因。

【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‘GanlüNo.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 en-zyme and key gene in the fruit of A.eriantha‘GanlüNo.1’at the ripening stage,A.eriantha‘GanlüNo.1’and another low sugar type kiwifruit cultivar,A.eriantha‘GanlüNo.6’,were selected as the re-search 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 Prov-ince.【Methods】The grafted vines of A.eriantha‘GanlüNo.1’and A.eriantha‘GanlüNo.6’were planted in an orchard of the Fengxin County Bureau of Agriculture and Rural Affairs,which was locat-ed in Fengxin County,Jiangxi Province.In 2021,at physiological maturity stage when the soluble sol-ids content of fruits of each cultivar reached over 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,the fruit weight was measured using an electronic balance;the fruit longitudinal diameter and fruit trans-verse diameter were measured respectively with a vernier caliper.The fruit shape index was obtained us-ing 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 mea-sured 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 de-termined by titration with NaOH;and the ascorbic acid(AsA)content was determined by the 2,6-di-chlorphenolindophenol titration method.The sugar-acid ratio was calculated by the ratio of the SS to the TA.The glucose(Glu),fructose(Fru),and sucrose(Suc)were also extracted from the frozen sam-ples in 80%ethanol,and their contents were determined by high-performance liquid chromatography.The total soluble sugar content(TS)was obtained by calculating the sum of Glu,Fru,and Suc.The sweetness value(SV)was calculated based on the contents of Glu,Fru,Suc,as well as their correspond-ing coefficients.Besides,for the frozen samples,the 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‘GanlüNo.1’and A.erian-tha‘GanlüNo.6’.In addition,the 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】The fruit size,fruit shape index,and fruit weight were signifi-cantly higher in A.eriantha‘GanlüNo.1’than those in A.eriantha‘GanlüNo.6’.Most of the indica-tors related to internal fruit quality were also significantly higher in A.eriantha‘GanlüNo.1’than those in A.eriantha‘GanlüNo.6’.Those indicators included SSC,SS,sugar-acid ratio,and AsA con-tent.Besides,higher contents of Fru,Suc,TS,and higher SV were detected in the fruits of A.eriantha‘GanlüNo.1’at the ripening stage.Additionally,in the fruits of GanlüNo.1,the activities of SPS and SUSY were both significantly higher than those of GanlüNo.6;however,the activities of AINV and NINV were significantly lower in the fruits of GanlüNo.1.Meanwhile,the relative expression levels of the AeSPS and AeNINV were significantly higher in the fruits of GanlüNo.1 than those of GanlüNo.6.Moreover,the 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 the Glu content,and the activi-ties of AINV and NINV.Furthermore,the relative expression level of the AeSPS was found to be posi-tively correlated to both the Suc content and SPS activity.【Conclusion】The difference in sucrose accu-mulation level was the dominant factor that results in the great difference in soluble sugar content be-tween the cultivars of A.eriantha‘GanlüNo.1’and A.eriantha‘GanlüNo.6’.And the sucrose phos-phate synthase was the key metabolic enzyme that could positively regulate sucrose accumulation in the ripe fruit of A.eriantha culitvars at the ripening stage.Besides,we speculated that the AeSPS would be the key gene which could contribute to the biosynthesis of sucrose by increasing SPS activity in the fruits of A.eriantha‘GanlüNo.1’at the ripening stage.