| 大豆rbcL基因克隆、序列分析及原核表达 |
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| 引用本文： | 刘晓庆,崔喜艳,丁志鑫,李海燕. 大豆rbcL基因克隆、序列分析及原核表达[J]. 中国油料, 2011, 0(3): 226-230 |
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| 作者姓名： | 刘晓庆 崔喜艳 丁志鑫 李海燕 |
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| 作者单位： | 吉林农业大学生命科学学院,吉林长春130118 |
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| 基金项目： | 国家转基因生物新品种培育重大专项（2008zx08010-002）;国家自然科学基金（30971804）;吉林省教育厅吉教科合字[2009]第61号 |
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| 摘 要： | 利用叶绿体基因组保守性的特征，根据菜豆、豌豆、烟草的rbcL基因序列设计引物，从大豆叶绿体DNA中克隆rbcL基因，全长序列为1488bp，包括1449bp的开放阅读框，编码482个氨基酸。相似性比较显示，此序列与其它10个物种rbcL基因核苷酸的同源性为85．37％～95．31％，氨基酸的同源性为90．87％-96．47％。将该基因与表达载体pET-30a（＋）连接，转化大肠杆菌Rosseta感受态细胞，PCR和酶切鉴定筛选阳性克隆，阳性菌液IPTG诱导后经10％SDS—PAGE分析，结果显示，诱导表达出分子量约为60kD的特异融合蛋白。
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| 关 键 词： | 大豆 rbcL基因 基因克隆 原核表达 |
Cloning, sequence analysis and prokaryotic expression of rbcL gene from Glycine max |
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| Affiliation: | LIU Xiao - qing, CUI Xi - yan, DING Zhi - xin, LI Hai - yan ( College of Life Sciences, Jilin Agricultural University, Changchun 130118, China) |
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| Abstract: | Rubisco is the key enzyme in plant photosynthesis. In this research, the large subunit of Rubisco, rbcL gene, from chloroplast of Glycine max was cloned and sequenced. Sequencing analysis indicated that this fragment was 1 488bp in size, including an ORF of 1 446bp, encoding a putative protein consisting of 482 amino acids. DNAman analysis results showed that the homologies of this gene with other species were from 85% to 96%, and the homologous amino acid sequences were from 90% to 97%. The rbcL gene was ligated into pET -30a（ ＋ ） and transformed into E. coli Rosseta competent cell. The positive clone was selected and sequenced. After induced IPTG, the fusion protein relative molecular weight was 60kD, which was consistent with the theoretical value. |
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| Keywords: | Glycine max rbcL gene Gene cloning Prokaryotic expression |
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