绿色棉苯丙氨酸解氨酶基因GhPAL1的克隆及实时定量表达分析.pdfVIP

分子植物育种，2012 年，第10 卷，第3 期，第311-316 页 Molecular Plant Breeding, 2012, Vol.10, No.3, 311-316 研究报告 Research Report 绿色棉苯丙氨酸解氨酶基因GhPAL1 的克隆和实时定量表达分析 1 2 2 1 2* 赵伟涛 李艳军 张新宇 吴莉 孙杰 1 石河子大学生命科学学院, 石河子, 832003; 2 石河子大学农学院, 新疆生产建设兵团绿洲生态农业重点实验室, 石河子, 832003 * 通讯作者, sunjiezh@ 摘 要 本研究以拟南芥(Arabidopsis thaliana)苯丙氨酸解氨酶(PAL)基因cDNA 序列(AAC18870.1)为探针， 在GenBank 中与棉花EST 进行tBLASTn 比对，将同源性高的EST 序列拼接后获得一条棉花PAL 全长 cDNA 基因序列，以此序列为模板设计引物，通过RT-PCR 扩增从绿色棉纤维分离出了一个绿色棉PAL 基 因全长cDNA 序列，将该基因命名为GhPAL1 (GenBank 登录号: JN032297)。序列分析表明，该cDNA 全长 2 347 bp ，含有一个编码721 个氨基酸的开放阅读框。实时荧光定量PCR 分析发现：该基因在绿色棉纤维中 的表达模式与白色棉纤维相似，但两者在同一时期的表达量差异显著，其在花后6 d 绿色棉纤维细胞中的表 达量是同期白色棉的6 倍以上，从该基因的表达特征推测GhPAL1 基因可能在绿色棉纤维色素合成过程中 发挥重要作用。 关键词 绿色棉, 苯丙氨酸解氨酶, 实时荧光定量PCR Cloning and Expression Analysis of a Phenylalanine Ammonia-lyase Gene GhPAL1 from Natural Green Cotton Fiber Zhao Weitao 1 Li Yanjun 2 Zhang Xinyu 2 Wu Li 1 Sun Jie 2* 1 College of Life Science, Shihezi University, Shihezi, 832003; 2 College of Agriculture, Shihezi University, The Key Oasis Eco-agriculture Laboratory of Xinjiang Production and Construction Group, Shihezi, 832003 * Corresponding author, sunjiezh@ DOI: 10.3969/mpb.010.000311 Abs tract In this study, the AtPAL amino acids sequence was used as a query sequence to make tBLASTn comparison with cotton ( Gossypium hirsutum ) EST database in GenBank. The highly homologous ESTs were identified, aligned and assembled into a full-length cDNA sequence, which was used to design specific primers. Then a new full-length cDNA sequence of PAL gene was isolated from green cotton fibers with RT-PCR method. This new gene was named GhPAL1 (GenBank accession No.JN032297). Sequence analysis reveals that its full-length cDNA Sequence is 2 347 bp, and has a open reading frame (ORF) encoding 721 amino acids. QRT-PCR analysis shows that the expressed pattern of GhPAL1 gene in green cotton fibers is similar to that in whi