银杏胞质抗坏血酸过氧化物酶基因(APX)克隆及表达特性分析(英文)

Cloning, characterization and expression analysis of a cytosolic ascorbate peroxidase gene (APX) from Ginkgo biloba

【目的】分离和克隆银杏细胞质抗坏血酸过氧化物酶基因GbAPX1(GenBank登录号为FJ555021),了解其在银杏正常组织发育以及响应环境胁迫和逆境相关激素诱导的表达情况。【方法】利用同源克隆及RACE技术从银杏叶片细胞质中分离得到GbAPX1全长cDNA序列,用生物信息学方法对获得的cDNA序列及推测的氨基酸序列进行分析,并用实时定量PCR方法研究GbAPX1在不同组织、不同激素和胁迫处理下的表达模式。【结果】克隆GbAPX1基因全长cDNA序列为965 bp,包含一个759 bp的最大读码框,编码252个氨基酸;预测该基因编码蛋白质分子量为27.6 kDa,其等电点为5.81。系统进化树分析表明,GbAPX1和其他裸子植物同源性较高并聚为一类。qRT-PCR分析结果显示,GbAPX1表达具有组织和发育时期差异性,在成熟叶片和雄蕊中具有最高表达水平,在茎和嫩叶中表达水平较低。诱导表达分析表明,GbAPX1受到激素和环境压力诱导。其中,脱落酸、水杨酸、茉莉酸甲酯和紫外辐射均能诱导其转录上升,而乙烯利和伤害处理对GbAPX1转录水平无显著影响。【结论】确认获得序列为GbAPX1基因cDNA序列;GbAPX1不仅参与了银杏正常的组织生长发育,还与一些环境胁迫和逆境相关激素诱导表达有关。

[Objective] The aim of this study was to isolate the APX homolog sequence from Ginkgo biloba L. （GbAPX1, GenBank accession No. FJ555021 ） and analyze its expression patterns in different tissues and in response to environmental stresses andstress-related hormones. [Methods ] Degenerate primer RT- PCR and RACE methods were used to clone GbAPX1. Bioinformatics methods were used to analyze its cDNA sequence and putative amino acid sequences, and real time quantitative PCR was used to study the expression of GbAPX1 in different tissues, under stresses and hormone treatments. [Result~ The full length cDNA of GbA PX1 was 965 bp, which contains an open reading frame of 759 bp, encoding a pro- tein of 252 amino acids with a predicted molecular weight of 27.6 kDa and an isoelectric point of 5.81. The phylogenetic analysis indicated that GbA PX1 was grouped together with A PXs in gymnosperm plants. The transcription of GbA PX] was tissue-specific and development-dependent, and the highest level was ob- served in mature leaves and stamens but only weakly expressed in young leaves and stems, qRT-PCRanalysis suggested that the expression of GbAPX1 gene was induced by both hormones and stresses. Its ex- pression level was increased by abscisic acid, salicylic acid, and methyl jasmonate treatments and by ul- traviolet, but ethephon and wounding had no significant effect on the expression of GbAPX1. [Conclu- sion] The studies suggested that GbAPX1 not only participates in the regulation of growth and develop- ment but also responses to stress-related hormones.