Analysis Of Dark Coloring Mechanism Of Bagged Apple Peel Induced By Light

Apples(Malus domestica Borkh.)belong to the deciduous fruit trees in the Rosaceae family,which are widely cultivated in temperate regions of the world.The color of apples is the main indicator of red-apple varieties,directly determining the appearance quality and commercial value of the fruits.The peel color is closely related to the biological metabolism of anthocyanins,and light is one of the important factors affecting the synthesis of fruit anthocyanins.Many studies have shown that strong light promotes the synthesis of apple anthocyanins,while dark or weak light weakens.The work discovered a phenomenon of light induction and coloring in the dark of apple peels.Spectral analysis showed that apples could synthesize anthocyanins normally at night.The key pigment substances,structural genes,and transcription factors for apples‘ coloring in the dark were screened out through metabolome and transcriptome analysis.Promoter analysis,yeast one-hybrid,EMSAand LUC experiments confirmed that MdNAC92 and MdLAF1 could positively regulate the transcription activity of MdUFGTs genes.The results of transient transformation experiment verified that MdLAF1,MdNAC92 and MdUFGTs all had positive regulation effect on anthocyanin synthesis.The main findings are as follows:1.This experiment studied the phenomenon of light induction and coloring in the dark of apples for the first time,that was,apple peels induced by a certain light could be colored normally in a completely dark environment.Simultaneously,spectroscopic analysis was used to verify that the light-induced peel produced anthocyanins at night on apples.In the process of light induction and coloring in the dark,bright chromaticity L*,comprehensive chromaticity h° and chlorophyll content decreased,while red chromaticity a* and anthocyanin content increased,with the fading green and red appearance on the apple-fruit surface.The stacked line graph of related enzyme activities showed that the key enzyme activity of anthocyanin synthesis was directly proportional to the duration of light induction.That was,the longer the light induction time,the greater the sum of related enzyme activities.2.Metabolome analysis detected 23 types of 620 metabolites.There were 121 flavonoid metabolites,including 10 anthocyanins,5 procyanidins,and 106 flavonoids(62flavonoids,30 flavonols,and 14 flavonoids).KEGG pathway enrichment showed that different metabolite enrichment between different treatments referred to flavonoid and anthocyanin biosynthesis pathways.Based on the test phenotype,cyanidin-3-O-galactoside,cyanidin-3-O-glucoside,anthocyanin,cyanidin-3-O-rutin,cyanidin-3-O-malonyl hexoside,O-hexoside,and pelargonium were screened out.Wherein,cyanidin-3-O-galactoside and cyanidin-3-O-glucoside are the two most important metabolites.3.Transcriptome analysis showed at most 2,467 difference gene CK?vs?G3 between treatments,of which 1,781 were up-regulated,and 686 were down-regulated.There were2,202 G3?vs?D7,of which 728 were up-regulated and 1,474 were down-regulated.KEGG pathway enrichment showed that anthocyanin biosynthesis was the key way for light induction and coloring in the dark in this experiment.Gene expression analysis showed that structural genes such as PAL,4CL,CHS,CHI,F3 H,F3'H,and ANS were light-sensitive.UFGT was the key structural gene,and MYB and NAC were the key transcription factors.Five MdUFGTs candidate genes,five MYBs transcription factors,and four NACs transcription factors were screened out by q RT-PCR verification.4.Promoter analysis showed that the promoter region of MdUFGTs had cis-acting elements that bind to MYB/NAC.The yeast one-hybrid experiment verified the interactions between MdNAC92 and MdUFGT,and that between MdLAF1 and MdUFGT.EMSA assay verified MdLAF1 binding to MdUFGT2 promoter and MdNAC92 binding to MdUFGT3 promoter.LUC experiments confirmed that MdNAC92 transcription activated the expression of MdUFGT3,and MdLAF1 transcription activated the expression of MdUFGT2.5.Instantaneous experiments on apple leaves showed that MdUFGTs promoted the biosynthesis of apple anthocyanins.Wherein,MdUFGT1 had the strongest promotion effect on anthocyanin accumulation,while MdUFGT3 and MdUFGT4 had relatively weak promotion effects.Transient expression of MdLAF1 and MdNAC92 in peel showed that MdLAF1 and MdNAC92 could positively regulate anthocyanin biosynthesis.