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百脉根钼转运蛋白基因LjMOT1的克隆与表达分析
发表时间：2017-02-16 浏览量：1893 下载量：407
| 全部作者： | 巫飞飞，沈知临，纪 萍，高俊山 |
| 作者单位： | 安徽农业大学生命科学学院 |
| 摘 要： | 克隆百脉根钼转运蛋白基因LjMOT1，其全长 cDNA 为1 824 bp，包括1 407 bp 的开放阅读框（open reading frame，ORF）、79 bp的 5¢-非编码区（untranslated region，UTR）和323 bp的3¢-UTR，以及15 bp的polyA，推测LjMOT1基因编码468个氨基酸。氨基酸序列比较显示百脉根LjMOT1蛋白与大豆GmMOT1蛋白的同源性最高，达到 81.14 %. 实时荧光定量PCR（quantitative real-time PCR，qRT-PCR）分析结果表明，百脉根根中LjMOT1的表达量随培养基中钼浓度的增加呈现出上升的趋势，而叶中在+Mo和 10×Mo 浓度下LjMOT1的表达量并没有明显的差异，但是含钼条件下叶中的LjMOT1表达量明显比-Mo条件下的高。构建真核表达载体pCambia1304-LjMOT1，在烟草上进行瞬时表达，通过激光共聚焦显微镜确定LjMOT1 定位于质膜上。这些结果为进一步鉴定LjMOT1的功能奠定了理论基础。 |
| 关 键 词： | 分子生物学；百脉根；钼；LjMOT1；实时荧光定量PCR；亚细胞定位 |
| Title： | Cloning and expression analysis of a molybdenum transporter gene LjMOT1 from Lotus japonicus |
| Author： | WU Feifei, SHEN Zhilin, JI Ping, GAO Junshan |
| Organization： | School of Life Sciences, Anhui Agricultural University |
| Abstract： | In this paper, a Mo transporter gene LjMOT1 was cloned from Lotus japonicus, and the full-length cDNA of LjMOT1 is 1 824 bp, which includes an 1 407 bp open reading frame (ORF), 79 bp 5¢-untranslated region (UTR) and 323 bp 3¢-UTR and 15 bp polyA. The LjMOT1 gene was predicted to encode 468 amino acids. The alignment of amino acid sequences showed the LjMOT1 protein had the highest identities (81.14%) with the GmMOT1 protein from soybean. The results of quantitative real-time PCR (qRT-PCR) illuminated that the expression level of LjMOT1 in roots appeared a rising trend with increasing Mo concentrations in the media. The expression level of LjMOT1 in leaves on +Mo and 10×Mo media had no significant difference, but they were obviously higher than those on -Mo medium. A eukaryotic expression vector pCambia1304-LjMOT1 was constructed, and was transiently expressed in tobacco. The result showed that the LjMOT1 protein was located on the plasma membrane by a laser scanning confocal microscope. These results provide theoretical basis for further identifying the function of LjMOT1. |
| Key words： | molecular biology; Lotus japonicus; molybdenum; LjMOT1; quantitative real-time PCR; subcellular localization |
| 发表期数： | 2017年2月第3期 |
| 引用格式： | 巫飞飞，沈知临，纪 萍，等. 百脉根钼转运蛋白基因LjMOT1的克隆与表达分析[J]. 中国科技论文在线精品论文，2017，10（3）：248-257. |
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