辣椒ASR基因的克隆与表达分析

【目的】克隆辣椒ASR（ABA-stress-ripening）基因进行生物信息学分析并探究其在不同组织和非生物胁迫处理下的表达模式，为深入研究ASR基因家族在辣椒果实发育和逆境胁迫中的作用机制提供参考。【方法】以遵辣1号辣椒为材料克隆其ASR基因，对该基因家族成员进行生物信息学分析和系统进化分析，运用实时荧光定量PCR检测其在不同组织及在ABA、高温、高盐和干旱胁迫处理下的表达模式。【结果】克隆的2个辣椒ASR基因分别命名为CaASR2和CaASR3，基因全长分别为987和938 bp，最长开放阅读框（ORF）分别为333和339 bp，分别编码110和112个氨基酸，蛋白分子量分别为12.45和12.73 kD，理论等电点（pI）分别为7.47和6.50，均为亲水性蛋白。结合前人克隆的辣椒CaASR1基因分析，结果显示，CaASRs预测位于细胞核，均含有ABA/WDS特征结构域；基因结构分析结果显示，辣椒ASR基因均含有2个外显子和1个内含子，启动子区域均包含ABA响应元件ABRE。CaASR1、CaASR2和CaASR3氨基酸序列分别与番茄（Solanum lycopersicum）基因组中的ASR蛋白氨基酸序列NP_001269248.1（80.20%）、NP_001307920.1（91.30%）和NP_001234137.1（96.43%）有非常高的相似性，表明其进化的相对保守性。3个ASR基因在辣椒花中表达量最低，在根、茎、叶和种子中表达量中等，但均随着果实的发育表达量逐渐升高，特别是在转色和成熟果实中。ABA、高温、高盐和干旱胁迫处理下，基因的表达量均显著升高（P<0.05），其中以CaASR3响应最快，且上调表达量高于其他同源基因。【结论】辣椒基因组中3个ASR基因具有ASR基因家族的典型特征，并在辣椒不同组织和非生物胁迫响应中发挥重要作用。

Cloning and expression analysis of ASR genes in pepper (Capsicum annuum L.)

【Objective】 ASR(ABA-stress-ripening) genes were cloned in this study. By bioinformatics analysis and expression analysis under different tissues and abiotic stress treatments, to provide a reference for further study on the mechanism of ASR gene family in pepper fruit development and abiotic stress.【Method】 ASR genes were cloned from Zunla-1 cultivar. A series of bioinformatics and phylogenetic analysis were carried out to evaluate the family members. The tissue specific expression patterns and the expression patterns under ABA, high temperature, high salt, and drought stress were detected by real-time fluorescence quantitative PCR.【Result】 Two ASR genes were cloned, named as CaASR2 and CaASR3. The full lengths of CaASR2 and CaASR3 were 987 and 938 bp, and the open reading frame(ORF) were 333 and 339 bp in length, which encoded 110 and 112 amino acids, respectively. Their molecular weights were 12.45 and 12.73 kD, and their theoretical isoelectric points(pI) were 7.47 and 6.50, respectively, and they were hydrophilic protein. The results showed that CaASRs were predicted to be in the nucleus and contained ABA/WDS(ABA/water deficit stress) domain. All CaASRs genes contained two exons and one intron, and the promoter region contained ABA response element ABRE. The amino acid sequences of CaASR1, CaASR2 and CaASR3 were highly similar to those of ASR protein amino acid NP_001269248.1(80.20%), NP_001307920.1(91.30%) and NP_001234137.1(96.43%) in tomato(Solanum lycopersicum), which indicated the relative conservation of evolution. The results of tissue specific expression analysis showed that they were expressed the lowest in flower, and the moderate in root, stem, leaf, and seed, gradually increased with fruit grew, and especially highly expressed in color-changing fruit and mature fruit. The expression levels were significantly increased under ABA stress, high temperature, high salt and drought(P<0.05), and CaASR3 had the fastest response, whose up regulated expression levels were higher than that of other homologous genes.【Conclusion】 Three ASR genes in pepper genome have typical characteristics of the ASR family and play important roles in different tissue and abiotic stress resistance in pepper.