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S-腺苷-L-高半胱氨酸水解酶(S-adenosyl-L-homocysteine hydrolase,SAHH)是已知的唯一能够水解S-腺苷高半胱氨酸(S-adenosylhomocysteine,SAH)成半胱氨酸和腺苷的酶。SAH作为转甲基反应的抑制剂,与S-腺苷甲硫氨酸(S-adenosyl-L-methionine,SAM,体内甲基供体)的比值常用来作为衡量植物整体甲基化水平的重要指标。为鉴定大叶落地生根(Kalanchoe daigremontiana)SAHH的功能,本研究根据已知的大叶落地生根SAHH基因(Kd SAHH)序列(Gen Bank登录号:KF953475)设计引物,对其进行干旱胁迫下的表达分析、亚细胞定位分析以及烟草(Nicotiana tabacum)转化,并对T1代苗用20%PEG6000模拟干旱处理,测定0、12和24 h的相对含水量(relative water content,RWC)、叶绿素(chlorophyll,Chl)含量、丙二醛(malondialdehyde,MDA)含量、过氧化物酶(peroxidase,POD)含量、超氧化物歧化酶(superoxide dismutase,SOD)含量等与抗旱相关的生理指标。结果表明,Kd SAHH基因在轻度(20 d)、中度(35 d)、重度(50 d)干旱胁迫下表达量依次升高,表明该基因与抗旱性相关;亚细胞定位结果显示,该蛋白定位于细胞质与细胞核;对转基因烟草进行PCR检测,获得9株阳性植株;测定干旱胁迫下的相关生理指标,结果显示,与野生型烟草相比,转Kd SAHH烟草在干旱胁迫下能够保持相对稳定的RWC、Chl含量以及相对较低的MDA、POD含量。与野生型相比,转Kd SAHH烟草受伤害轻,抗旱性增强,表明Kd SAHH基因能够提高烟草植株的抗旱性。研究结果丰富了Kd SAHH基因的功能,为后续研究该基因提供了借鉴和参考。
S-adenosyl-L-homocysteine hydrolase (SAHH) is the only enzyme known to hydrolyze S-adenosylhomocysteine (SAH) Cysteine and adenosine enzyme. SAH, as an inhibitor of transmethylation reactions, is commonly used as a measure of the overall methylation level in plants as a ratio of S-adenosyl-L-methionine (SAM, in vivo methyl donor) index. In order to identify the function of SAHH in Kalanchoe daigremontiana, primers were designed according to the known sequence of Kd SAHH (GenBank accession number: KF953475) and under drought stress Analysis, subcellular localization analysis and Nicotiana tabacum transformation. The relative water content (RWC), chlorophyll Chl content, malondialdehyde (MDA) content, peroxidase (POD) content, superoxide dismutase (SOD) content and other drought-related physiological indicators. The results showed that the expression level of Kd-SAHH gene increased in turn under light (20 d), medium (35 d) and severe (50 d) drought stress, indicating that the gene was related to drought resistance. The results of subcellular localization The protein was localized in the cytoplasm and the nucleus. Nine transgenic plants were obtained by PCR. The relative physiological indexes under drought stress were determined. The results showed that the transgenic Kd-SAHH tobacco could maintain the relative drought resistance under drought stress Stable RWC, Chl content and relatively low MDA, POD content. Compared with the wild type, Kd SAHH tobacco was harmed lightly and drought resistance was enhanced, indicating that the Kd SAHH gene can improve the drought resistance of tobacco plants. The results enrich the function of Kd SAHH gene and provide references for further study of this gene.