大麦穗发育候选基因 HνAP2–260 的原核表达载体构建及其编码蛋白质制备

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中图分类号：S512.303.2 文献标志码：A 文章编号：1004-390X（2025）06-0001-07

Abstract： [Purpose] To construct the prokaryotic expression vector of candidate gene HvAP2-260 in barley young spike development， and to prepare its high purity encoding protein. [Methods] The specific PCR primers with sequence recognized by enzyme NdeI and XhoI were used to amplified cDNA templates， from young spike of barley variety Morex at jointing stage. Multi-stage recombination method and directed linking methods were employed to sequentially construct the recombinants pET- 30a（+）-HνAP2-260 and pCZN1-HvAP2-260; then， heat shock was used to transfer the recombinants into competent cels of Escherichia coli strain Trans1-Tl and Arctic Express，respectively.By using E . coli transformed with pCZN1 empty vector and uninduced positive clones as controls， the expression differences of protein encoded by the gene HνAP2–260 under different treatments were analyzed， and the expressed proteins were purified by affinity chromatography. [Results] The 924 bp coding sequence （CDS） of gene HνAP2–260 wascloned successfully， a prokaryotic expression vector pCZN1-HvAP2-260 was constructed. The protein encoded by the HνAP2–260 gene could be efficiently induced to express in the Arctic-Express strain cells，but the expressed proteins were inclusion bodies.The expressed proteins could be effectively purified by using a Ni-IDA-Sepharose Cl-6B chromatography column. [Conclusion] The CDS of the barley spike development gene HνAP2–260 （20号 has been successfully cloned， the prokaryotic expression vector has been constructed， and the highpurity proteins encoded by the gene have been prepared. The prepared proteins are useful for followup studies， such as antibody preparation， functional verification and action mechanism elucidation of the protein encoded by gene HvAP2-260.

Keywords： HordeumvulgareL.; HνAP2–260 gene clone; expression vector construction; prokaryotic expression; protein preparation

在长期进化过程中，栽培大麦（Hordeumvul-gareL.）形成了特有的花序结构，即：由中间花序轴（也称为"穗轴"和两侧三联小穗（由3个并列单花小穗构成）组成，花序轴无分枝，三联小穗直接着生在花序轴节上1。（剩余12033字）