谷子类受体蛋白激酶基因SiRLK35的克隆及原核表达.doc

谷子类受体蛋白激酶基因SiRLK35的克隆及原核表达 　　摘要：本研究以“豫谷1号”cDNA为模板，通过PCR技术获得类受体蛋白激酶基因SiRLK35（NCBI登录号：XM_004956247.2 ）的全长序列。生物信息学分析显示，该基因编码蛋白由392个氨基酸残基构成，包含一个S_TKc保守结构域以及一个TFS2N结构域，含3个跨膜结构域，N端有信号肽。 SiRLK35基因经BamHⅠ、SalⅠ双酶切后，利用pET-28a构建原核表达载体，转化大肠杆菌菌株BL21（DE3），经0.5 mmol/L IPTG于25℃诱导12 h后，在44.6 kD处得到一明显诱导表达蛋白条带，表达产物与预期相符，证明SiRLK35基因可在大肠杆菌中诱导表达。该结果为进一步研究SiRLK35基因功能奠定基础。 　　关键词：谷子；类受体蛋白激酶；SiRLK35；原核表达 　　中图分类号：S515+Q78文献标识号：A文章编号：1001-4942（2016）09-0001-05 　　Cloning of Receptor-Like Protein Kinase Gene SiRLK35 　　from Foxtail Millet and Its Prokaryotic Expression 　　Wang Yifan1， 2，Li Zhen1，Pan Jiaowen1，Wang Qingguo1，Liu Wei1* 　　（1. Biotechnology Research Center， Shandong Academy of Agricultural Sciences/Shandong Provincial Key Laboratory of 　　Crop Genetic Improvement， Ecology and Physiology， Jinan 250100， China； 　　2. College of Life Sciences， Qingdao Agricultural University， Qingdao 266109， China） 　　AbstractIn this research， the full-length sequence of receptor-like protein kinase gene SiRLK35 （NCBI accession number： XM_004956247.2） was amplified by PCR technique with the cDNA of Yugu 1 as template. Bioinformatics analysis showed that the protein product of SiRLK35 was composed of 392 amino acids. It contained one S_TKc conserved domain， one TFS2N domain， three transmembrane domains and a N terminal signal peptide. The sequence of SiRLK35 was digested with BamHⅠ and SalⅠ， and then the recombined prokaryotic expression vector pET-28a-SiRLK35 was constructed successfully through connecting the fragment with pET-28a. The recombinant was further transformed into E.coli strain BL21 （DE3）. Induced with 0.5 mmol/L IPTG， a 44.6 kD fusion protein was obtained， which proved that the SiRLK35 could be induced to express in E.coli. This research layed a good foundation for further study of the function of SiRLK35 gene. 　　KeywordsFoxtail millet； Receptor-like protein kinase； SiRLK35； Prokaryotic expression 　　谷子[Setaria italica （L.） Beauv.]又称为粟，是起源于我国的古老作物，具有抗旱耐贫瘠的特点，是旱作生态农业的重要组成部分。近年来，其营养保健价