Cloning And Analysis Of MYB Transcription Factors Gene Related To Lignin Synthesis In Postharvest Wax Apple Fruit

Wax apple(Syzygium samarangenese [Blume] Merrill & L.M.Perry)fruit has bright color,rich nutrition and unique flavor.After harvesting,due to the thin skin and cottony softening of the fruit,the cottony softening in the center of the fruit will continue to expand to the outside,which will seriously affect the taste and flavor.To explore the role of transcription factors in Lignin Metabolism of Postharvest wax apple fruits is helpful to understand the regulation mechanism of flocculent deterioration of wax apple fruits,but there are few reports.This study selects Taiwan honey campanula wax apple fruit as experimental materials,to research Nitric oxide(NO)treatment on the physiological quality and lignin synthesis of postharvest wax apple fruits.Twenty MYB transcription factors related to lignin synthesis were screened from the wax apple transcriptome database,selecting the SsMYB1 gene with the largest expression difference was cloned and analyzed for subcellular location.In order to explain the effect of MYB transcription factors on the lignin metabolism of the postharvest wax apple fruit.The main results of this study are as follows:1.Wax apple treated with the concentration of 10 ?L/L and 0 ?L/L(control)NO on the harvested wax apple.The results showed that during the storage period(0-12 d),the weight loss rate and cottony softening index of Postharvest wax apple fruit increased,the flesh firmness of pulp decreased gradually,and the soluble solid content and total sugar content increased first and then decreased;the activities of key enzymes PAL,C4 H,4CL,CAD and POD showed an upward trend.Compared with the control group,the NO treatment group slowed down the increase in fruit weight loss rate,the expansion of cottony softening index,the decrease of fruit firmness,the decrease of soluble solids and total sugar content,inhibition of lignin synthesis and key enzyme activity.Therefore,thereby delaying fruit lignification and maintaining better fruit quality.2.20 MYB transcription factors related to lignin synthesis that were screened from the wax apple transcriptome database established in the early stage of the research group,Name the members of the SsMYBs family according to the result of species homology comparison,which are: SsMYB1,SsMYB3,SsMYB4,SsMYB5,SsMYB6,SsMYB10,SsMYB20,SsMYB33,SsMYB39,SsMYB44,SsMYB48,SsMYB,SsMYB48,SsMYB,SsMYB54,SsMYB77,SsMYB77 SsMYB306,SsMYB308,SsMYB330 and SsMYBC1.Bioinformatics analysis was performed on these 20 MYB transcription factors.The results showed that the 20 MYB proteins related to lignin synthesis in wax apple are composed of 91-591 amino acids,the relative molecular mass of the corresponding protein is 9.98-64.91 k D,and the theoretical isoelectric point(p I)is 4.92-9.73,mostly unstable hydrophilic protein,no signal peptide,subcellular localization on the nucleus,no obvious transmembrane domain,and not a secreted protein.According to phylogenetic tree analysis,wax apple is closely related to Syzygium oleosum,and Eucalyptus grandis,and Rhodamnia argentea.The expression analysis results during the wax apple fruit storage period showed that the expression levels of SsMYB1,SsMYB4,SsMYB20,SsMYB54,SsMYB77,SsMYB108,SsMYB114 and SsMYB308 showed an overall upward trend,and NO treatment slowed the upward trend.It is speculated that it may be a transcription activator to promote the synthesis of lignin,While the expression of SsMYB3,SsMYB5,SsMYB6,SsMYB10,SsMYB33,SsMYB39,SsMYB44,SsMYB48,SsMYB86,SsMYB306,SsMYB330 and SsMYBC1 decreased during the storage period.NO treatment suppressed this downward trend,and it is speculated that they may be transcription repressors,delaying the synthesis of lignin.This result further shows that the selected 20 wax apple MYB transcription factors play a regulatory role in the postharvest lignin synthesis of wax apple fruit.3.Select the SsMYB1 gene,which is most obviously expressed during storage,was selected for cloning,the full-length sequence was 1230 bp,and the subcellular localization of the SsMYB1 gene was verified.The plant expression vector p CAMBIA1301-35 SN and green fluorescent protein(GFP)were fused and expressed by the fusion gene location method.The recombinant expression plasmid SsMYB1-EGFP-p CAMBIA1301-35 SN was constructed,and subcellular localization was performed by Agrobacterium infection method.The distribution of recombinant plasmid in onion epidermal cells was observed by fluorescence microscope to determine the location and expression of SsMYB1 gene in the cell.The research results showed that the recombinant expression vector of the SsMYB1 gene is located on the nucleus,indicating that the SsMYB1 protein is a nuclear protein.