从加州鲈(Micropterus salmoides)脑组织中克隆获得NF-κB家族中p65 基因的编码区序列,该基因cDNA全长 1 944 bp,ORF为1 899 bp,可编码632个氨基酸.理论分子质量为69.61 ku,理论等电点(PI)为 5.66,通过生信学分析工具对p65 基因蛋白结构进行分析,同时对其蛋白功能进行预测,结果显示:该基因不含有信号肽序列,同时不存在跨膜结构域,但存在1 个NF-κB家族保守的结构域.多序列比对结果显示,加州鲈(Micropterus salmoides)p65与小口黑鲈(Micropter us dolomieu)的相似度最高,达99.30%;构建系统进化树发现,加州鲈(Micropterus salmoides)p65与小口黑鲈(Micropter us dolomieu)聚为一支.通过qRT-PCR结果表明,加州鲈p65 基因在所取组织均有表达,在脑组织中表达量最高,其次是头肾、胸腺和心脏,在脾脏组织中的表达量最低.
Cloning and Expression Analysis of p65 Gene in Micropterus salmoides
Cloning and obtaining the coding region sequence of the p65 gene in the NF-κ B family from the brain tissue of Micropterus salmo-ides,with a total cDNA length of 1 944 bp,the ORF is 1 899 bp and can encode 632 amino acids.The theoretical molecular weight is 69.61 ku,and the theoretical isoelectric point(PI)is 5.66.The protein structure of the p65 gene was analyzed using bioinformatics analysis tools,and its protein function was predicted.The results showed that the gene does not contain a signal peptide sequence and does not have a transmembrane domain,but there is a conserved domain in the NF-κ B family.The results of multiple sequence alignment show that the simi-larity between Micropterus salmoides p65 and Micropterus dolomieu is the highest,at 99.30%;constructing a phylogenetic tree,it was discov-ered that the Micropterus salmoides p65 and Micropterus dolomieu clustered together.The results of qRT PCR indicate that the Micropterus salmo-ides p65 gene is expressed in all selected tissues,with the highest expression level in brain tissue,followed by the head kidney,thymus,and heart,and the lowest expression level in spleen tissue.
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