青杄PwPSAF基因的克隆与组织表达分析

doi:10.11707/j.1001-7488.20131007

摘要/Abstract

摘要：

PSAF是绿色植物光系统Ⅰ反应中心F亚基,在光合作用电子迁移过程中起着重要作用。以青杄cDNA文库为模板,根据PwPSAF的EST序列设计引物,进行5'-RACE和3'-RACE试验,以获得青杄PwPSAF的末端序列,经过与EST序列进行拼接获取青杄PwPSAF的全长cDNA序列。基于其氨基酸序列,利用Expasy tools、DNAMAN、ClustalX和MEGA等工具,进行理化性质、二级结构和三级结构的生物信息学分析。结果表明: PwPSAF是一个由253 aa组成的蛋白质,理论分子量为27.04 kDa,等电点为9.57,预测蛋白有2个跨膜区域,蛋白质的C端具有保守的结构。RT-qPCR试验显示PwPSAF主要在青杄的针叶中表达。在种子萌发过程中吸胀期(0~2天)表达量最高,在4天时表达量降低,随着子叶的不断长出,其表达量又逐渐增高。在干旱和盐胁迫下,针叶中的PwPSAF的表达量被大幅下调。研究结果显示PwPSAF在青杄种子萌发及非生物逆境胁迫下的光合过程中起着重要作用。

关键词: 青杄, PSAF, 生物信息学分析, 组织表达, 胁迫响应

Abstract:

PSAF is F subunit of photosystem Ⅰ reaction center complex in higher plants and plays an important role during the process of electron mobility of photosynthesis. The full length cDNA of PwPSAF was obtained by RACE PCR assays based on the cDNA library of Picea wilsonii and EST fragment of PwPSAF. Several pieces of software and tools such as Expasy, DNAMAN, ClustalX and MEGA were used to analyze the physical and chemical properties and secondary and tertiary structures of PwPSAF. Expression level of PwPSAF in different tissues of seedlings was identified by RT-qPCR under drought and salt treatments. Furthermore, the expression patterns were investigated at different stages of seed germination. We found that PwPSAF was composed of 253 amino acids and the theoretical molecular weight was 27.04 kDa with isoelectric point of 9.57. The protein has two transmembrane domains and shares conserved C domain with other species such as Picea sitchensis and Vitis vinifera. RT-qPCR assays indicated that PwPSAF was mainly expressed in needles of Picea wilsonii. During seed germination, PwPSAF showed the highest expression within 2 days after germination, and then declined on the fourth day. With the cotyledons grown, the expression of PwPSAF was up-regulated again. In addition, the expression level of PwPSAF was down-regulated in the needles under drought and salt stresses. These results suggest that PwPSAF in P. wilsonii plays an important role during seed germination and in response to abiotic stresses.

Key words: Picea wilsonii, PSAF, bioinformatic analysis, tissue-specific expression, stress response

中图分类号:

S718.46

李长江, 孙帆, 张通, 张凌云. 青杄PwPSAF基因的克隆与组织表达分析[J]. 林业科学, 2013, 49(10): 40-47.

Li Changjiang, Sun Fan, Zhang Tong, Zhang Lingyun. Cloning and Tissue Expression Analysis of PwPSAF in Picea wilsonii[J]. Scientia Silvae Sinicae, 2013, 49(10): 40-47.

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