基于氮素对枸杞甜菜碱含量及代谢关键酶影响的转录组分析

Transcriptome analysis based on effect of nitrogen on betaine content and key metabolic enzymes in Lycium barbarum

【目的】探究不同施氮量对枸杞主要次生物质甜菜碱及合成关键酶含量基因水平的影响,为宁夏枸杞品质代栽培提供技术支持。【方法】以‘宁杞1号’为试验材料,运用裂区试验设计,主区为氮肥施用深度A的2个处理(A1:0~20 cm,A2:20~40 cm),副区为氮素施用量B的5个处理(CK:0 kg/666.7 m^(2),B1:15 kg/666.7 m^(2),B2:30 kg/666.7 m^(2),B3:45 kg/666.7 m^(2),B4:60 kg/666.7 m^(2)),通过枸杞生长期间相关指标测定及不同氮素水平枸杞样品高通量下的转录组测序分析,明确氮素对枸杞甜菜碱及代谢关键酶的影响。【结果】随着施氮量的增加,枸杞甜菜碱及其合成相关酶含量基本呈上升趋势,在浓度达45 kg/666.7 m^(2)时,所测定指标含量开始下降。转录组测序共获得98264个基因,其中差异表达基因10081个。GO注释分析表明,注释基因主要富集在光合作用、质外体、单加氧酶活性、叶绿素、四吡咯结合酶活性等过程中;进行KEGG注释后,分析发现差异基因信号通路显著富集在植物激素信号传导、植物病原菌互作以及核糖体中;对与甜菜碱合成相关酶差异基因进行分析,发现其表达量均有不同程度的上调。综合分析所测定指标含量和基因表达水平之间的关系,表明甜菜碱合成相关酶基因的上调,导致枸杞果实内甜菜碱含量发生变化。【结论】甜菜碱合成关键酶是单加氧酶和甜菜碱醛脱氢酶,随着施氮量的适量增加,甜菜碱合成关键酶含量及其相关基因数量上调,致使枸杞果实内甜菜碱含量上升。

【Objective】The present paper aimed to explore the effects of different nitrogen application rates on the gene levels of betaine and key enzyme contents in Lycium barbarum,and provide technical support for the quality generation cultivation of L.barbarum in Ningxia.【Method】In the experiment,‘Ningqi No.1’was used as the experimental material,and the split⁃plot experiment design was used.The main area was two treatments of nitrogen fertilizer application depth A(A1:0-20 cm,A2:20-40 cm),and the sub⁃area was five treat⁃ments of nitrogen application amount B(CK:0 kg/666.7 m^(2),B1:15 kg/666.7 m^(2),B2:30 kg/666.7 m^(2),B3:45 kg/666.7 m^(2),B4:60 kg/666.7 m^(2)).Through the determination of related indicators during the growth period of L.barbarum and the high⁃throughput tran⁃scriptome sequencing analysis of L.barbarum samples with different nitrogen levels,the effects of nitrogen on betaine and key metabolic en⁃zymes in L.barbarum were clarified.【Result】With the increase of nitrogen application rate,the contents of betaine and its synthesis⁃related enzymes in L.barbarum were basically on the rise.When the concentration reached 45 kg/666.7 m^(2),the contents of the measured indexes began to decrease.A total of 98264 genes were obtained by transcriptome sequencing,including 10081 differentially expressed genes.GO annotation analysis showed that the annotated genes were mainly enriched in photosynthesis,apoplast,monooxygenase activity,chlorophyll,tetrapyrrole binding enzyme activity and other processes.After KEGG annotation,it was found that the differential gene signal patnway was significantly enriched in plant hormone signal transduction,plant pathogen interaction and ribosome.The differentially expressed genes related to betaine synthesis were analyzed,and it was found that the expression levels were up⁃regulated to varying degrees.Comprehensive anal⁃ysis of the relationship between the content of the measured indicators and the results of gene expression levels showed that the up⁃regulation of betaine synthesis⁃related enzyme genes led to changes in betaine content in L.barbarum fruits.【Conclusion】The key enzymes of betaine synthesis are monooxygenase and betaine aldehyde dehydrogenase.With the increase of nitrogen application rate,the content of key enzymes in betaine synthesis and the number of related genes are up⁃regulated,resulting in the increase of betaine content in L.barbarum fruit.