半夏凝集素基因的克隆与氨基酸序列初步分析-天津中草药杂志社.pdfVIP

• 1818 • 中草药 Chinese Traditional and Herbal Drugs 第 43 卷 第 9 期 2012 年 9 月 半夏凝集素基因的克隆与氨基酸序列初步分析 张正英 甘肃省农业科学院，甘肃 兰州 730070 摘 要：目的 克隆半夏凝集素基因并对其氨基酸序列生物信息与已报道相关基因进行对比分析，为抗虫基因利用和转基因 育种研究奠定基础。方法 根据已报道的植物凝集素基因序列设计特异引物，以半夏叶片 DNA 为模板进行 PCR 扩增，获 得特异性片段，将其连接到测序载体上，进行序列测定，用分析软件分析序列信息。结果 克隆 1 069 bp 的半夏凝集素基因 4 (pta ) ，开放阅读框全长 804 bp，编码268 个氨基酸残基；预测相对分子质量和等电点分别为 2.91 ×10 和 7.77，功能区完整， 具有 1 条信号肽和 3 个甘露糖结合区；已在 GenBank 中登记（登录号AY725425 ）。结论 克隆的半夏凝集素基因序列信息 完整，具有典型的甘露糖结合位点，可以作为抗虫基因用于抗虫育种。 关键词：半夏；半夏凝集素（PTA ）；基因克隆；序列分析；特异性片段 中图分类号：R282.12 文献标志码：A 文章编号：0253 - 2670(2012)09 - 1818 - 06 Cloning of Pinellia ternata agglutinin gene and analysis on its amino acid sequence ZHANG Zheng-ying Gansu Academy of Agricultural Sciences, Lanzhou 730070, China Abstract: Objective To clone Pinellia ternata agglutinin (PTA) gene, compare its amino acid sequence and bioinformation with former reported genes, and lay the foundation for utilization of insect-resistant gene and transgenic breeding. Methods Based on the specific primers designed by reported plant lectin gene sequence, DNA extracted from P. ternata leaves was taken as template to amplify PCR and a specific fragment was obtained which was linked to the sequencing vector for being sequenced and their information was analyzed using analytical software. Results A full-length nucleotide sequence of pta with 1 069 bp was cloned (AY725425 in Genbank). The full length of open reading frame (ORF) was 804 bp encoding 268 amino acids residue. The predicted relative molecular weight and isoelectric point (PI) were 2.91 × 104 and 7.77, respectively. The functional section was intact with a signal peptide and three mannosebinding sites. Conclusion The cloned pta gene is intact