Study On The Sepal Shedding Of ’Yuluxiangli’ Based On The Analysis Of GH3 Gene Family

Whether pear sepals fall off or not is an important factor affecting the formation of fruit quality.Spraying paclobutrazol can improve the calyx dropping rate and fruit quality of pear fruits,but there is no report about its mechanism in the sepals off of pear fruits.In this experiment,compared the fruit quality difference with the detached and permanent calyx fruits which was measured with the ’Yuluxiang Pear’cultivated on a trellis,analyzed the change of the IAA content in the calyx under different chemical treatments and its relationship with the peeling of pear’s sepals,combined with transcriptome sequencing data,analyzed the spatiotemporal expression pattern of GH3 family of auxin amide synthase genes in pears,cloned GH3 candidate members and performed bioinformatics analysis,to reveal the molecular mechanism of pear sepal shedding or persistence and propose new research ideas.There are main findings of this article :1.The ‘Yuluxiang Pear’ cultivated in a trellis type used as the experimental material,we studied the differences in the appearance quality,internal quality and texture of the ‘Yuluxiang Pear’ detached calyx fruit and persistent calyx fruit in the mature period.The results showed that the single fruit weight of detached calyx was significantly lower than that persistent calyx fruit,and the fruit shape index was close to 1.The soluble solid content of detached calyx fruit was significantly higher than that of persistent calyx,and the titratable acid content and water content were significantly lower than that of persistent calyx;the flesh firmness and gelatinity are significantly lower than that of persistent calyx fruits.The detached calyx fruit has medium size,the fruit shape is more suitable,and the flavor is better.2.Enzyme-linked immunoassay measured the IAA content in the calyx of the detached calyx fruit and the persistent calyx fruit,it was found that the IAA content of the detached calyx was lower than that of the persistent calyx.Spraed paclobutrazol,gibberellin and water treatment in the early blooming stage,the relationship between the calyx removal rate at the end of full blooming and the change of the IAA content showed that as the IAA content of the calyx part gradually decreased,the calyx removal rate of’Yuluxiangli’ increased.Analyzd the differences in gene expression of the auxin signaling pathway revealed that three members of the GH3 family of auxin amide synthase genes were highly expressed in the detached calyx.3.Analyzed the GH3 gene family,there are 18 members in the GH3 gene family in Pyrus bretschneideri.The isoelectric point PI is between 5.23 to 9.13,the molecular weight is between15.26-69.58(KD),and the largest number of chromosomes distributed on Chr5.The subcellular locations are mainly distributed in the cytoplasm,plasma membrane and mitochondrial matrix.A phylogenetic tree was constructed with the Arabidopsis GH3 protein,it was found that the pear GH3 protein are closely related to the Arabidopsis I and II proteins.The 9 relatively conservative motif types indicate that the GH3 protein family is highly conserved in pears;all 18 GH3 family genes have predicted cis-acting elements related to hormone regulation.Ten genes including Pb GH3.2,Pb GH3.3,Pb GH3.7,Pb GH3.8,Pb GH3.9,Pb GH3.10,Pb GH3.13,Pb GH3.16,Pb GH3.17 and Pb GH3.18 have auxin response elements(Aux RR-core and TGA-element).4.To further study the expression pattern of GH3 gene family in ’Yuluxiang Pear’,the fluorescence quantitative analysis was performed on different parts of pears and the calyx tube of paclobutrazol-treated.The expression levels of Pb GH3.2,Pb GH3.3,Pb GH3.5,Pb GH3.10,Pb GH3.11,Pb GH3.15,Pb GH3.16,Pb GH3.17 and Pb GH3.18 in sepals were higher than that in other parts;under paclobutrazol treatment,Pb GH3.2,Pb GH3.3,Pb GH3.4,Pb GH3.5 and Pb GH3.10 expression levels were the highest in the late full bloom,but were significantly different from the control group.5.Cloned Pbr007550(Pb GH3.3)and Pbr030571(Pb GH3.4),both of which are 1806 bp in length.However,there are differences in three amino acid sequences,encoding 601 amino acids.They belong to unstable protein,without signal peptide,and subcellular localization in cytoplasm.Using the NCBI website CCD tool analysis found that the GH3 protein family has the GH3 super famliy conserved domain,protein three-level modeling found it to be GH3-1 AUXIN conjugating enzyme,involving in auxin homeostasis,belonging to the GH3 protein family.