Study On Physiological And Molecular Response Mechanism Of Low Temperature And Low Light Resistance Of Capsicum Pepper Based On Exogenous Melatonin

Low temperature and low light are common environmental stress in the off season vegetable production of protected horticulture,which may occur in the whole growth period of plants.Pepper is the main cultivation crop for off-season vegetable production in facilities.In 2018,the domestic pepper planting area has exceeded 32 million mu,accounting for 21.4%of the world's pepper planting area,and 9.3%of the national vegetable planting area.However,the growth and development of pepper are vulnerable to low temperature and low light stress,resulting in the decline of yield and quality.Therefore,it is of great significance to improve the low temperature and low light resistance of pepper to ensure the sustainable production of pepper in China.In this study,low temperature and low light conditions(15?/5?,100 mol.m^-2.s^-1)were simulated under normal growth conditions and artificial climate chamber,and melatonin(200 mol.L^-1)was sprayed from external sources.The physiological and biochemical responses of melatonin on growth index,chlorophyll fluorescence,photosynthetic performance,osmoregulation substance,antioxidant enzyme and glutathione cycle system to low temperature and low light stress were studied under four treatments:CK,CK+M,LL and LL+M.Transcriptomic sequencing was used to analyze the molecular mechanism of melatonin in relieving low temperature and low light stress of capsicum,which providing a theoretical basis for the application of exogenous melatonin in relieving stress in the production of capsicum.The main conclusions are as follows:(1)Through the observation of the external morphology of pepper seedlings,after 24 hours,the leaves of LL treatment were obviously wilted,but the leaves of LL+M treatment were not serious.After 7 days of treatment,compared with LL treatment,LL+M treatment had significantly higher plant stem diameter,leaf number,aboveground dry and fresh weight and underground dry and fresh weight,transpiration rate increased by 288.71%,stomatal conductance increased by 102.75%,net photosynthetic rate increased by 254.17%,and intercellular CO₂concentration decreased by 21.66%.Exogenous application of melatonin could effectively alleviate the damage of chilling and low light stress on the growth and photosynthetic capacity of pepper plants.(2)The contents of chlorophyll a,b,a+b and total carotenoids in LL treatment were significantly lower than those in CK treatment,indicating that low temperature and low light inhibited the chlorophyll content of leaves significantly.The contents of chlorophyll a,b,a+b and total carotenoids in LL+M treatment were significantly higher than those in LL treatment,indicating that exogenous melatonin alleviated the effect of low temperature and low light on the decrease of chlorophyll content.(3)Through the chlorophyll fluorescence imaging of leaves,it can be clearly seen that the damage degree of the four treatments is different after 7 days.The indexes of Fo,Fm and Fv/Fm show that LL treatment is the most damaged,LL+M treatment is slightly injured,and CK+M treatment is not significantly different from CK.The results showed that the low temperature and low light stress significantly reduced the photosynthetic capacity of pepper plants;the q L,q P,Y?,ETR of LL+M treatment were significantly higher than those of LL treatment,and q N and NPQ of LL+M treatment were significantly lower than those of LL treatment,indicating that melatonin could alleviate the damage effect of low temperature and low light stress and effectively improve the chlorophyll fluorescence parameters of leaves.(4)According to the analysis of growth indicators,there was no significant difference between CK and CK+M treatment on chlorophyll fluorescence,photosynthetic performance,osmotic regulatory substances,antioxidant enzymes,endogenous hormones,glutathione circulation system and other indicators.Therefore,exogenous melatonin had no significant influence on the growth and development of capsaicin under normal growth conditions(P<0.05).(5)Based on physiological and biochemical characteristics of early test results,the transcriptome analysis set up the control(CK),low light and low temperature(LL),low temperature and low light+melatonin(LL+M)three treatments.The leaves of pepper seedlings at different time points were sampled and transcriptome sequencing was performed on Illumina Hiseq4000 platform.The analysis results showed that total RNA was extracted from the leaves of pepper seedlings treated with CK,LL and LL+M at 0h,6h,12h,24h,48h and 168h,and 54 c DNA libraries were constructed.A total of 2,380,892,332 raw reads were obtained,and 350.68G high quality sequence was obtained by sequencing.The remaining 2,337,948,108 clean reads were filtered,in which,the Q20?97.5%and the Q30?91.37%;the GC content was between 42.26%and 43.48%.(6)Compared with LL,LL+M treated at 0h,6h,12h,24h,48h and 168h had 4666(upregulated 2332,downregulated 2334),4865(upregulated 2450,downregulated 2415),2260(upregulated 1276,downregulated 964),1981(upregulated 841,downregulated 1140),4511(upregulated 2415,downregulated 2096)and 1435(upregulated 891,downregulated 544),respectively.They were significantly enriched in 14,42,12,14,13,26 GO Terms and 2,15,4,10,2,2 KEGG pathways,respectively.In which 14 differences related to the photosynthesis genes are expressed are raised,article 9 of the gene encoding the PS?reaction center,four genes encode PSI reaction center;Carbon metabolism in 31 differentially expressed genes related raised expression has 15,16 cut expression,two of the four genes related to sucrose metabolism were raised to express,and dextran and glucose metabolism in seven of nine genes increase express two lower expression,protein synthesis and hydrolase in five genes associated with a higher expression of four lower expression,and galacturonic acid phosphate enzyme activity related 6 2increase in gene expression,four lower expression,three genes associated with wood glycosidase activity all cut expression;Among the 6 differentially expressed genes related to glucose metabolism,2 were up-regulated and 4 were down-regulated;among the 5 genes related to hydrolase protein synthesis,XTH10 was up-regulated,XTH22,XTH30,LOC107864353 and XTH28 were down-regulated,and the CSLG2 gene related to cellulose protein synthesis was up-regulated.Antioxidant activity of the 13 differentially expressed genes associated with eight raised expression,5 lower expression,10 genes associated with the activity of peroxidase in 7raised expression,3 lower expression,related to the types of cystic cavity of protein synthesis CLEB3J9 raised expressed genes,SODA genes associated with superoxide dismutase(SOD)expression of NADPH oxidase activity related RBOHE genes expression.(7)After melatonin treatment,the differential expression of genes in antioxidant metabolic pathways such as photosynthesis,carbon metabolism,starch and sugar metabolism,hormone metabolism,glutathione cycle and ascorbic acid cycle played an important role in alleviating low temperature and low light stress in capsaicin.The specific biological and molecular mechanisms need to be further studied.