| Polygonatum cyrtonema Hua is a perennial herb of the genus Polygonatum in the family Liliaceae,and its rhizome is used medicinally as a traditional medicinal herb of the same origin.The rhizome mainly contains polysaccharides,flavonoids,saponins and a variety of amino acids that are beneficial to the human body,and has the effect of tonifying the qi and nourishing the yin,strengthening the spleen,moistening the lungs and benefiting the kidneys,etc.Light regulates all aspects of plant morphogenesis,physiology,biochemistry and metabolism.Light environment exists in the whole growth cycle of plants and is constantly changing.Light is also one of the important environmental factors affecting plant growth and development and accumulation of metabolic substances.In this study,the effects of different light conditions including different photoperiods,light intensities and light qualities on the growth of Polygonatum cyrtonema Hua seedlings and rhizomes,as well as the contents of polysaccharides,flavonoids and saponins,the main metabolites of Polygonatum cyrtonema Hua rhizomes,were investigated to provide a theoretical reference for the in vitro fast breeding and factory production of Polygonatum cyrtonema Hua.The gene family of Polygonatum cyrtonema HuaACY1 was identified and expressed,and the overexpression vector of Polygonatum cyrtonema Hua ACY1 was constructed to transiently transform the rhizomes of Polygonatum cyrtonema Hua and analyze the expression of PcACY1 under different light conditions.The main findings of the study were as follows:1 Effect of different light conditions on the growth of seedlings and rhizomes of Polygonatum cyrtonema HuaThe results of different photoperiodic treatments on the growth of Polygonatum cyrtonema Hua seedlings showed that 6 h/18 h light duration was relatively the most suitable light duration for the growth of seedlings and rhizomes of Polygonatum cyrtonema Hua.The growth indexes measured all showed a decreasing trend with the increase of light time and showed a maximum at 6 h/18 h or 12 h/12 h,and the rhizome proliferation coefficient and weight gain coefficients were significantly higher in the 6 h/18 h treatment than in the other treatments.The results of different light intensities showed that the light intensity of 6000-8000 lx significantly promoted the proliferation of seedlings and rhizome weight gain of Polygonatum cyrtonema Hua;the results of different light qualities of Polygonatum cyrtonema Hua seedlings showed that single green light had a strong inhibitory effect on the growth of seedlings and rhizome of Polygonatum cyrtonema Hua,while blue light significantly inhibited plant height but promoted the proliferation of rhizome shoots and the accumulation of photosynthetic pigments in the leaves of Polygonatum cyrtonema Hua seedlings,while red light promoted the growth of leaves in Polygonatum cyrtonema Hua seedlings.2 Effect of different light conditions on the main metabolites of rhizomes of Polygonatum cyrtonema HuaThe results of different photoperiodic treatments showed that the 6 h/18 h photoperiod treatment promoted the rapid accumulation of polysaccharides,while the 18 h/6 h long photoperiod treatment promoted the accumulation of flavonoids and saponins in the rhizomes of Polygonatum cyrtonema Hua;the results of different light intensity treatments showed that the contents of Polygonatum cyrtonema Hua.The results of different light intensities showed that the polysaccharide,flavonoid,and saponin contents were basically positively correlated with light intensity,and the highest values were observed in the high light intensity treatment of 6000-8000 lx.The results of different light quality treatments showed that the accumulation of polysaccharide and saponin contents was inhibited by a single light quality treatment for more than 30 days,and this inhibitory effect would deepen with time,but blue light could promote the accumulation of flavonoids in the rhizomes of Polygonatum cyrtonema Hua.3 Identification and expression analysis of the ACY1 family members of Polygonatum cyrtonema HuaTwo PcACY1 members,named PcACY1-1 and PcACY1-2,were identified from the full-length transcriptome of Polygonatum cyrtonema Hua by bioinformatics,with sequence lengths of 1371 bp and 1317 bp in the CDS region,respectively,and isoelectric points of 5.35 and 5.42 for the two members,indicating that they mainly function in a weakly acidic cellular environment and are both hydrophilic proteins with a signal peptide but no transmembrane structure.The phylogenetic tree was constructed by screening 65 ACY1 members from 32 species from the plant genome database.The analysis revealed that most plant species have two members of the ACY1 family,indicating that the number of ACY1 genes in these species is relatively constant and the evolution of ACY1 in plants is highly conserved.qPCR detected that the PcACY1 gene is not present in different tissue sites under salt stress or at high temperatures.The expression of the PcACY1 gene in different tissue sites under salt stress and high temperature stress was found to be higher in rhizomes and the highest in stalks,indicating that the expression of PcACY1 members is tissue specific and that all PcACY1 family members were induced by salt stress and high temperature treatments,suggesting that members of the ACY1 family of Polygonatum cyrtonema Hua play an important role in stress resistance and defense.The PcACY1 family members were all induced by salt stress and high temperature treatment,indicating that the ACY1 family members of Polygonatum cyrtonema Hua play an important role in stress resistance and defense balance.4 qPCR analysis of PcACY1 in different light conditionsThe expression of PcACY1 family members differed under different light conditions;the expression of PcACY1-1 may not respond to photoperiodic changes,while the expression of PcACY1-2 responded to a 12h/12h light time treatment;the expression of PcACY1-1 was significantly up-regulated in all light intensity treatments,while the expression of PcACY1-2 was significantly up-regulated in all light intensity treatments.The expression of PcACY1-1 was significantly downregulated in all light intensity treatments,whereas the expression of PcACY1-2 was significantly upregulated in response to high light intensity treatments of 6000-8000 lx.PcACY1-1 significantly responded to blue light treatments,whereas the expression of PcACY1-2 may not be induced by a single light quality such as red,blue,or green light.5 Construction and transient transformation of ACY1s overexpression vector from Polygonatum cyrtonema HuaSuccessful construction of pCAMBIA1 301-PcACY1-1 and pCAMBIA1301-PcACY1-2 overexpression vectors and successfully transiently transformed overexpressed PcACY1-1 and PcACY1-2 into the rhizomes of Polygonatum cyrtonema Hua seedlings using an Agrobacterium-mediated method.In addition,the expression of PcACYl1 and PcACY1-2 genes in the rhizomes of Polygonatum cyrtonema Hua was analyzed by real-time fluorescence quantitative PCR,and it was found that the expression of the genes in the transient transgenic rhizomes of Polygonatum cyrtonema Hua was significantly higher than that of the wild type,but the expression decreased under different light quality treatments.In summary,the short photoperiod and high light intensity promoted the growth and rhizome metabolite accumulation of Polygonatum cyrtonema Hua seedlings,while blue light in a single light quality inhibited the plant highly,but promoted rhizome proliferation and flavonoid accumulation.Two ACY1 members were identified in Polygonatum cyrtonema Hua with tissue-specific expression patterns and played an important role in stress resistance and defence homeostasis,and the successful construction of the overexpression vector of Polygonatum cyrtonema Hua ACY1s also laid the foundation for further in-depth study of their functions. |
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