白灵侧耳ACC基因克隆及表达分析  

Cloning and Expression Analysis of ACC Gene in Pleurotus tuoliensis

作　　者：田雨 郝刘斌 李海康 冯璠 王春霞 张瑞颖[2] 郑素月 TIAN Yu;HAO Liubin;LI Haikang;FENG Fan;WANG Chunxia;ZHANG Ruiying;ZHENG Suyue(College of Landscape and Ecological Engineering,Hebei University of Engineering,Handan 056038,Hebei,China;Institute of Agricultural Resources and Agricultural Zoning,Chinese Academy of Agricultural Sciences,Beijing 100081,China)

机构地区：[1]河北工程大学园林与生态工程学院,河北邯郸056038 [2]中国农业科学院农业资源与农业区划研究所,北京100081

出　　处：《食用菌学报》2024年第5期9-16,共8页Acta Edulis Fungi

基　　金：国家现代农业产业技术体系(CARS-20);河北省现代农业产业技术体系创新团队建设(HBCT2023090207)。

摘　　要：通过克隆白灵侧耳(Pleurotus tuoliensis)乙酰辅酶A羧化酶(acetyl-CoA carboxylase,ACC)基因,采用生物信息学方法预测白灵侧耳ACC的理化性质、信号肽、跨膜结构域、亚细胞定位、二级结构、三级结构和保守结构域;基于白灵侧耳ACC与灰盖鬼伞(Coprinopsis cinerea)、糙皮侧耳(P.ostreatus)、香菇(Lentinula edodes)、刺芹侧耳(P.eryngii)等16个物种的氨基酸序列,采用邻接法构建系统发育树;通过荧光定量PCR测定pH为3、4、5、6、7、8、9、10、11、12时的白灵侧耳ACC的相对表达量。结果表明:白灵侧耳ACC DNA长度为7366 bp,包含12个内含子和13个外显子,cDNA长度为6696 bp,共编码2231个氨基酸残基,相对分子质量为247400,白灵侧耳ACC为酸性、不稳定的亲水蛋白质,不具有信号肽和跨膜结构域,可能存在于细胞质、细胞核或过氧化物酶体中,主要由α螺旋、无规卷曲、β折叠和β转角组成,含有ACC中心区、生物素羧化酶、生物素羧基载体蛋白质和羧基转移酶4个保守结构域;系统发育树表明,白灵侧耳ACC的氨基酸序列与担子菌中的刺芹侧耳相似度较高;与对照组(pH5.42)相比,pH为6、7、8、9、10、11时,ACC相对表达量显著增加(P<0.05)。The acetyl-CoA carboxylase(ACC) gene of Pleurotus tuoliensis(PtACC) was cloned,and the encoded ACC protein was analyzed by bioinformatics tools for physicochemical properties,signal peptide,transmembrane structure,subcellular localization,secondary structure,tertiary structure and conserved domains.Based on the ACC protein sequence of P.tuoliensis(PtACC) and that of other 16 species,such as Coprinopsis cinerea,P.ostreatus,Lentinula edodes,and P.eryngii,a phylogenetic tree was constructed using the neighbor-joining algorithm.The relative expression of PtACC gene under different pH(3,4,5,6,7,8,9,10,11,and 12) was measured by fluorescence quantitative PCR.The results showed that the DNA sequence of PtACC was 7 366 bp,comprising 12 introns and 13 exons,and its cDNA was 6 696 bp,encoding a protein product of 2 231 amino acids with a molecular weight of 247.4.The PtACC protein was predicted to be an unstable acidic hydrophilic protein without any signal peptide or transmembrane structure,and is likely to present in the cytoplasm,nucleus or peroxisome.It is primarily composed of α-helix,random coil,β-sheet and β-turn.It has four conserved domains,ACC_central,AccC(BC),AccB(BCCP),and carboxyl_trans(CT).Phylogenetic analysis showed that PtACC was culstered with the ACC of P.eryngii.Compared with the control pH of 5.42,the relative expression of PtACC was significantly higher at pH 6,7,8,9,10 and 11.

关 键 词：白灵侧耳 乙酰辅酶A羧化酶 基因克隆 系统发育 表达 

分 类 号：S646[农业科学—蔬菜学]