The Research On The Mechanism Of Red Light Transmission Bagging To Improve Carotenoids Accumulation In The Flesh Of Blood Orange Fruit

As the largest fruit over the world.Citrus is widely loved by consumers because of its rich color and nutritional value.Bagging is a common agronomic measure to improve fruit quality.Light is not only an important environmental factor affecting fruit quality formation,but also the most important factor for bagging treatment to change fruit growth conditions.Therefore,the light changes caused by bagging are the most important factor in the formation of fruit quality with this technique,but the mechanism is unclear.Carotenoids,as the main pigment and functional nutrients,are closely related to the appearance quality and nutritional value of citrus fruits.In this study,by bagging’Moro’ blood orange fruit,it was found that the content of total carotenoids in the flesh of ’Moro’ blood orange fruit was significantly increased by red light transmission bagging treatment;Then,the molecular mechanism of ’Moro’ blood orange flesh mediating red light quality signal regulating carotenoid accumulation was analyzed by using transcriptome sequencing and citrus fruit transient expression technology.The main results are as follows:1.A total of 50 different carotenoids were identified from the flesh of ’Moro’ blood orange by liquid chromatography tandem mass spectrometry(LC-MS / MS)technology,including 4 carotenes,namely phytoene,α-Carotene,β-Carotene,γ-Carotene;12 luteins,namely α-Cryptoxanthin,β-Cryptoxanthin,lutein,zeaxanthin,antherxanthin,rubixanthin,violaxanthin,neochrome,neoxanthin,apocarotenal,astaxanthin,β-citraurin.Among them,violaxanthin and phytoene are the main components of carotenoids in ’Moro’ blood orange flesh,accounting for 84.76% or more.Under different light bagging transmission treatments,the composition of carotenoids is essentially the same in blood orange flesh,but the proportion of each composition and content had a similar change trend during ripening.Among them,the proportion of violaxanthin and lutein decreased at the mature stage(DAB273,i.e.273 days after blossom),and the rest showed an increasing trend.2.In this study,seven luteins,namely β-Cryptoxanthin,lutein,zeaxanthin,antheraxanthin,rubixanthin,violaxanthin and neochrome,were identified from the flesh of ’Moro’ blood orange,which can combine with different fatty acids to form a total of37 carotenoid esters,including 13 monoesters,24 diesters.Esterified carotenoids are the main form of storage in the flesh of blood orange.During the color transition period(DAB210),the total content of esterified carotenoids accounts for 82.12% of the total content of carotenoids.In the mature stage(DAB273),although it decreased,it still accounted for 60.04% or more.3.Carotenoids in the flesh of ’Moro’ blood orange accumulated continuously during fruit ripening.The total carotenoid content in the flesh of ’Moro’ blood orange reached 352.41 ～ 472.54 μg/g at the mature stage(DAB273),which is 3.94 ～ 4.18 times of the commercial harvest time(DAB210).Compared with the control group,the red light transmission bagging treatment significantly induced the accumulation of carotenoids in both periods.Violaxanthin-myristate and violaxanthin-myristate-caprate are the main contributors to carotenoid accumulation induced by red light transmission bagging treatment.4.RNA-seq analysis revealed that red light transmission bagging treatment significantly induced the expression of carotenoid pathway genes.Among them,it promoted the expression of the synthetic genes PDS(102616616),ZDS(102616177),CRITISO(102577949),ZEP-like(112497075),the phytochrome genes PHYA(102616503),PHYB(102618185),PHYC(102618478).and suppressed the expression of the cleavage genes CCD1-like(102615521),CCD4-8(102621234),NCED2(102578070)and NCED3(102578064),which may be involved in metabolic pathways associated with the regulation carotenoid metabolism by red light transmitting bagging treatment.5.The expression of red light receptor phytochrome genes was detected to be clearly changed in the red light transmissive bagging treatment,and it was hypothesized that PHY family genes may be key genes in regulating this process.In this study,Cs PHYA2 was subsequently identified as an important candidate gene based on genome-wide family analysis and gene expression analysis.To further validate its function,overexpression of Cs PHYA2 was found to significantly promote not only carotenoid accumulation but also the expression of the synthetic genes CRITISO(102577949)and ZEP-like(112497075)and suppress the expression of the cleavage gene NCED2(102578070)through citrus fruit transient expression,confirming that Cs PHYA2 promotes the accumulation of carotenoids by specifically regulating genes in their metabolism.