Correlation And Mechanism Of Starch Metabolism And Physicochemical Quality In Apple Fruit During Postharvest Based On RNA-Seq

Apple(Malus domestica)is a woody plant of the Rosaceae Malus genus,and is one of the important fruits widely grown at home and abroad.Most apple fruits have post-ripening phenomenon after harvest.However,the understanding of the symbolic changes of their optimal eating period is very insufficient,which seriously limits the income-increasing potential of high-quality and high-price apples and the quality of apple consumption.Based on the research group’s earlier finding that different varieties of apples will degrade slowly at a certain level during the storage time of different varieties of apples.‘Granny Smith’,‘Jonagold’,‘Golden Delicious’,‘Qinguan’,’Fuji’ and ’Ruiyang’ were used as test materials to explore the correlation between starch changes and other physicochemical indexes during postharvest storage in low-temperature,and to analyze the fruit physicochemical quality at key points of postharvest starch changes;RNA-Seq technology was used to sequence the transcriptome of 6 varieties of apple pulp at different storage time,and the differentially expressed genes enriched in the starch and sucrose metabolism pathways were mined.Through the quantitative verification of gene expression and the determination of enzyme activity,the starch metabolism of postharvest apples was analyzed.The preliminary molecular mechanism of the turning point is expected to provide a theoretical and technical basis for establishing a method for determining the optimal eating period of apples based on the key node of starch change.The main results are as follows.(1)There are key points of starch change in all six varieties of apples during low temperature storage.At this time,the starch content is between 0.20～0.26%.The storage periods for ’Granny Smith’,’Jonagold’,’Golden Delicious’,’Qinguan’,’Fuji’ and ’Ruiyang’ to reach the key node of starch change were 80,40,60,30,30,and 60 d,respectively,and the comprehensive evaluation of the six apple varieties reached the best at the key node of starch change.(2)During low temperature storage,the soluble sugar content,reducing sugar content and starch content of 6 varieties of apples were significantly correlated;the titratable acid content of ’Jonagold’,’Qinguan’ and ’Fuji’ was extremely significantly correlated with starch content,which of ’Granny Smith’,’Golden Delicious’ and ’Ruiyang’ was significant correlated with starch;the correlation of firmness and starch among ’Granny Smith’,’Jonagold’,’Golden Delicious’ and ’Qin Guan’ was extremely significant,which was significant between ’Fuji’ and ’Ruiyang’ was significantly correlated with starch content.Except ’Ruiyang’,the content of soluble sugar and titratable acid of the other 5 varieties all reached the equilibrium point near the key node of starch change(crossover of relative values),and the comprehensive evaluation of apple quality of 6 varieties was in the change of starch key node reaches the best.(3)Compared with the storage period of 160 d and the storage period of 0 d,there were1503 DEGs that were significantly differentially expressed in the pulp of the six varieties of apples,and a total of 156 DEGs were enriched in the KEGG pathway of starch and sucrose metabolism,involving 26 species of enzymes.Among them,there are 5 enzymes related to starch synthesis and degradation,including 31 DEGs,as well 6 enzymes related to sucrose synthesis and degradation,including 21 DEGs.Meanwhile,11 genes related to starch degradation were screened,MdISA1,MdISA2,MdISA2 L,MdSBE1,MdSEBL,MdAMY3,MdAMY10,MdBAM,MdBAM2,MdPHSL1,MdPHSH1.(4)The gene members of isoamylase(ISA),MdISA1 and MdISA2,involved in starch decomposition,reached the maximum expression levels in ’Jonagold’ and ’Fuji’ at 30 or 40 days of storage.The expression level of two members of α-amylase(AMY),MdAMY3 and MdAMY10,reached the maximum at 60 days,and the expression levels of MdAMY3 and MdAMY10 in ’Jonagold’ and ’Fuji’ reached the maximum at 30 or 40 days after storage,and were significantly higher than those at 0 d,which increased by 80-200 times,and reached the maximum at 60 d in ’Granny Smith’,and increased by 100-160 times compared with 0 d.The two members of β-amylase(BAM),MdBAM and MdBAM2,both reached the maximum value at 60 d,and were 2-25 times higher than that of 0 d;Pho-L starch phosphorylase(starch phosphorylase L isozyme,Pho-L SP)gene member MdPHSL1 was continuously upregulated,and starch branching enzyme(SBE)gene member MdSBE1 and MdSEBL involved in starch synthesis,Pho-H starch phosphorylase(starch phosphorylase H isozyme,Pho-H SP)gene member MdPHS1 was continuously down-regulated.(5)The AMY activities of ’Granny Smith’,’Jonagold’,’Golden Delicious’,’Qinguan’,’Fuji’ and ’Ruiyang’ reached 60,40,40,30,30,and 30 d storage periods,respectively.BAM activity peaked at 60 d.The neutral invertase(NI),acid invertase(NI),and sucrose synthase cleavage(SUS-c)involved in the degradation of sucrose in the fruits of the 6 varieties were all critical for starch degradation.It gradually increases before the node and maintains a higher level later.The activities of sucrose synthase synthesis(SUS-s)and sucrosephosphate synthase(SPS)involved in sucrose synthesis were also enhanced after key nodes.The SPS activities of 3 middle maturity varieties and 3 late maturity varieties also reached their peaks at 60 d and 90 d,respectively.In this paper,the uniform value range of starch content,which marks the best physical and chemical quality of 6 varieties postharvest apples,ranges from 0.20% to 0.26%.And also revealed the periodic changes,physiological and partial molecular mechanisms of starch and soluble sugar in apple fruits after harvest,and identified MdAMY3,MdAMY10,MdBAM and MdBAM2 as key genes for rapid starch degradation.The characteristic indicators of preservation provide the basis for selecting the optimal consumption period of apples and the characteristic indicators of apple quality preservation.