| Kiwifruit is climacteric fruit,with hard texture and acid taste before mature,while the flavor and taste of kiwifruit reach the optimum after full mature.Unlike other varieties with inferior preservation properties,’Hayward’ kiwifruit can be stored longer,although the preservation mechanism remains poorly reported.Consequently,the author sequenced Small RNAs of ’Hayward’ kiwifruit on three periods(DAP 1,DAP 15 and DAP24,respectively),compared the sequence results with Gene-Bank database and Mir-database,and ascertained the target gene of Small RNAs and analyzed their function accordingly.To test the sequence results,the expressions of six miRNA involved in ethane metabolism,IAA metabolism and ABA metabolism of kiwifruit were determined.Additionally,ethephon,ABA and 1-MCP were adopted as interference reagent to conduct symbiotic stress experiment to further investigate the effects of the six miRNA on endogenous hormones.The results are as follows:1)Sequence results of Small RNAs on DAP1,DAP15 and DAP24The reads amounts of Small RNAs on DAP1,DAP15 and DAP24 were 11489447,11995421,10800862,respectively.145 known miRNA were detected and 576 unknown miRNA were found on DAP1.On DAP 15 and DAP 24,137 and 164 known miRNA and 522 and 1739 unknown miRNA were respectively detected.Expression of miRNA was strongest on DAP1 and decreased with the prolonged period.These findings suggested that miRNA had significant effects on kiwifruits during post-mature period.2)prediction results of target genes of miRNARobot and Rangefinder software were used to predict the target genes of miRNA and results showed that 1108 target genes were detected on DAP1,followed by DAP 15(1175 target genes),and DAP24(3149 target genes).Besides,576 unknown miRNA regulating 2896 genes,522 unknown MiRNA regulating 3176 target genes and 1739 miRNA regulating 8871 genes were detected on DAP1,DAP15,DAP24,respectively.3)verification of expression of miRNAqRT-PCR analysis showed that the sequence results of 6 miRNA involved in ethane metabolism,IAA metabolism and ABA metabolism,were in accordance with expression trend,proving the reliability of the sequence results.4)effects of ethephon,ABA and 1-MCP on expression of 6 miRNA’Hayward’ kiwifruit was treated with ethephon,ABA and 1-MCP,respectively,and hardness value,soluble solid content and thane content and six miRNA content of treated fruits were determined regularly during the storage.The results showed that ethephon and ABA increased the release of thane,and 1-MCP retarded the decrease of fruit hardness and the presence of thane release peak.Besides,ethephon increased the expression of miR162a-3p,miR172c-3p and miR8129,whereas inhibited the expression of miR7545,miR9484,and miR5158.For fruits treated with ABA,the expression of miR162a-3p and miR9484 significantly increased(p<0.01),while the expression of miR172c-3p decreased.The expression of miR162a-3p,miR7545 and miR9484 significantly increased(p<0.01)during the storage whereas the expression of miR172c-3p decreased.Additionally,1-MCP increased the expression of miR8129 and decreased the expression of miR172c-3p,miR7545 and miR5158 in varying degrees.All results suggested that miRNA expression were closely related with the post-mature of kiwifruit,revealing the metabolism of internal hormone resulted from interaction of those miRNA.5)expression of PYR/PYL,PP2C and ABF(the target genes of miR9484 and miR7545)To study the effects of miRNA on ABA anabolism,’Hayward’ kiwifruits were treated with ethephon,ABA and 1-MCP,respectively,and the expression of key genes involved in ABA metabolism pathway were determined.Results suggested that 1-MCP significantly increased(p<0.01)the expression of PYR/PYZ(Achn125031),PP2C(Achn236131)and ABF(Achn016621)during post-mature period.Compared with control group,ABA increased the expression of ABF,PYR and PYL,decreased the expression of PP2C.What’s more,ethephon stimulated the expression of PYR/PYL during middle and last storage,whereas decreased the expression of PP2C and ABF during the same time. |
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