低温对青海扁茎早熟禾活性氧代谢的影响和相关基因表达分析

Effect of freezing stress on ROS metabolism and cold-induced genes in Poa pratensis var. anceps cv. Qinghai

为研究青海扁茎早熟禾(BJ)在低温胁迫下活性氧积累、抗氧化酶防御与逆境相关基因表达的机制,以WY-2(草地早熟禾‘午夜2号')为对照,对这两种材料低温适应(10/5℃)3 d和冷冻胁迫(-10℃)24 h,测定了O_2^(-·)产生速率、H_2O_2含量、丙二醛(MDA)含量、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)、抗坏血酸过氧化物酶(APX)和谷胱甘肽还原酶(GR)活性及其抗氧化酶相关基因、冷胁迫相关转录因子的相对表达量。结果表明,低温胁迫可引起BJ和WY-2叶片内的O_2^(-·)产生速率、H2O2含量和MDA含量的增加,BJ活性氧的积累程度和MDA含量均低于WY-2;低温胁迫可导致BJ的SOD、APX和GR活性的降低,CAT活性呈先升后降的趋势,但只有APX活性表现为BJ高于WY-2,同时,BJ和WY-2叶片内的Cu/Zn SOD、CAT、APX和GR基因的表达量在低温胁迫后被明显的上调,且BJ抗氧化酶相关基因的相对表达量普遍高于WY-2;BJ叶片内的DREB2是下调表达的,ERF、DREB1、CSP和HSP基因在低温胁迫后均表现出明显的上调表达,且ERF、CSP和HSP的相对表达量高于WY-2。从而得出结论,扁茎早熟禾在低温胁迫时有害物质的积累低于午夜二号,抗氧化酶相关基因表达上调幅度,以及一些与冷胁迫相关转录因子的表达调控均大于午夜二号。这些物质合成和基因表达的差异可能共同导致扁茎早熟禾的高耐冷性。

To explore the mechanism of cold tolerance of Poa pratensis var. anceps cv. Qinghai（ BJ） plants were exposed to cold acclimation（10/5 ℃） for 3 d followed by freezing stress（-10 ℃） for 24 h,with P. pratensis var.‘Midnight II＇（WY-2） as control,and ROS accumulation,antioxidant enzyme activity and stress related gene expression were monitored. Rate of the production of O2^-·,levels of H2O2 and MDA,activity of SOD,CAT,APX,GR and the relative expression of the antioxidant enzyme genes were measured,as well as several cold induced genes. The results demonstrated that freezing stress increased the O2^-·production rate,and H2O2 and MDA levels in both BJ and WY-2.ROS accumulation and MDA content of BJ were lower than those in WY-2. SOD,APX and GR activity was depressed by freezing stress in BJ,while CAT activity initially increased and then decreased. Only APX in BJ showed higher activity than that in WY-2. At the same time,the expression levels of Cu/Zn SOD,CAT,APX and GR related genes were increased in the two varieties. The relative expression of the enzyme related genes in BJ was higher than WY-2. The cold induced gene DREB2 was down-regulated in BJ,while the ERF,DREB1,CSP,and HSP genes were up-regulated. Among these genes,ERF,CSP and HSP showed higher expression levels in BJ than in WY-2. In conclusion,the accumulation of ROS in BJ was lower than that in WY-2 under cold and freezing stress. Meanwhile antioxidant enzyme related genes and cold related transcription factors showed higher relative expression levels in BJ than in WY-2 under cold and freezing stress. Those differences may contribute to the higher cold tolerance of BJ.