摘要
目的克隆珍稀濒危兰科药用植物铁皮石斛蛋白磷酸酶(protein phosphatase,PP)基因DoPP2C2并进行生物信息学和表达模式分析。方法采用RACE和RT-PCR获基因全长;利用生物信息学软件预测蛋白的理化性质、结构域和亚细胞定位等分子特性;用DNASTAR 7.0和MEGA 6.0分别进行氨基酸多序列比对和进化关系分析;借助定量PCR检测基因表达模式。结果分离到DoPP2C2基因(GenBank注册号KT957553),cDNA全长1624 bp,编码一条由387个氨基酸组成的多肽,相对分子质量42.76×103,等电点7.03;DoPP2C2蛋白具有植物PP2Cs蛋白家族保守的2个蛋白磷酸酶结构域(91~152、216~376);该蛋白无信号肽或跨膜域,定位在细胞核,系亲水性蛋白;DoPP2C2蛋白与多种植物PP2C蛋白一致性较高(47.8%~73.4%);DoPP2C2隶属于拟南芥和水稻PP2C蛋白家族进化树的A亚族,与OsPP2C30亲缘关系较近;DoPP2C2基因表达有器官差异,根和茎中相对表达量为叶的0.76和0.59倍。结论获得一个A亚族铁皮石斛蛋白磷酸酶DoPP2C2基因全长及分子特征,为深入研究该基因在铁皮石斛生长发育中的分子作用奠定基础。
OBJECTIVE To characterize a protein phosphatase(PP)encoding gene DoPP2C2 in a rare endangered medicinal orchid species Dendrobium officinale,followed by bioinformatics analysis and expression profiling.METHODSE RACE and RT-PCR were used to isolate the full length cDNA of DoPP2C2.The physiochemical properties,conserved domains and subcellular localization of the deduced DoPP2C2 protein were determined using a series of bioinformatics tools.The analyses of multiple alignment and phylogenetic tree were performed using DNASTAR 7.0 and MEGA 6.0 softwares,respectively.Quantitative PCR was used for gene expression analysis.RESULTS The full length cDNA of DoPP2C2(GenBank accession KT957553)was 1624 bp in length,and encoded a 387-aa protein with a molecular weight of 42.76×103 and an isoelectric point of 7.03.The deduced DoPP2C2 protein sequence had two PP2C domains(91-152,216-376),which are all conserved among the PP2Cs.The protein without a signal peptide or a transmembrane region,was predicted to locate in nucleus with hydropathicity.DoPP2C2 had high identities(47.8%-73.4%)with various PP2C proteins from several plants.DoPP2C2 protein belonged to the subgroup A of Arabidopsis and rice PP2C evolutionary tree,and was closely related to OsPP2C30;DoPP2C2 was differentially expressed in the three included organs.The transcripts were less in the roots and stems,being 0.76 and 0.59 folds,respectively,of that in the leaves.CONCLUSION The molecular characteristics of a subgroup protein phosphatase encoding gene DoPP2C2 of the full length cDNA in D.officinale was obtained.The results will be useful for further functional determination of the gene involving in the growth and development of D.officinale.
作者
李依民
李元敏
梁小燕
张明英
高静
颜永刚
郭顺星
张岗
LI Yi-min;LI Yuan-min;LIANG Xiao-yan;ZHANG Ming-ying;GAO Jing;YAN Yong-gang;GUO Shun-xing;ZHANG Gang(College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine,Shaanxi University of Chinese Medicine,Xi′an 712046,China;Institute of Medicinal Plant Development,Chinese Academy of Medical Sciences-Peking Union Medical College,Beijing 100193,China)
出处
《中国药学杂志》
CAS
CSCD
北大核心
2020年第14期1195-1200,共6页
Chinese Pharmaceutical Journal
基金
陕西省自然科学基金项目资助(2019JQ-872)
陕西中医药大学思邈青年学者人才项目资助(141306200105)
陕西中医药大学“秦药”品质评价及资源开发学科创新团队项目资助(2019-QN01)
陕西中医药大学新进博士科研启动经费资助(104080001)
中国医学科学院医学与健康科技创新工程经费资助(2017-I2M-3-013)
陕西省高校青年杰出人才支持计划项目资助
咸阳市中青年科技领军人才项目资助。
关键词
铁皮石斛
蛋白磷酸酶
生物信息学
定量PCR
信号
Dendrobium officinale
protein phosphatase
bioinformatics analysis
quantitative PCR
signaling
