摘要
从砀山酥梨Pyrus bretschneideri ‘Dangshan Su’果实中克隆得到一条多酚氧化酶基因(PbPPO)的全长cDNA序列(GenBank登录号:JF809859)。通过生物信息学分析表明,PbPPO基因CDS区全长1782 bp,编码593个氨基酸,氨基酸序列由N端叶绿体转运肽、Cu结合区与C端疏水区三部分组成。为进一步研究Pb PPO基因的功能,成功构建其原核表达载体pET-28a-PbPPO,并在大肠杆菌Rosetta菌株中成功诱导表达,经优化后显示,在28℃、1.0 mmol·L^(-1) IPTG诱导5 h的条件下,融合蛋白表达量最高。
This study cloned the full-length cDNA sequence of a polyphenol oxidase gene （PbPPO） from Dangshan Su pear. The bioinformatics analysis showed that the full-length ofPbPPO gene was 1782 bp, encoding 593 amino acids. The amino acid sequence consisted of three parts： N terminal chloroplast transport peptide, Cu binding region and C-terminal hydrophobic region. In order to further study the fimction of PbPPO gene, the prokaryotic expression vector pET-28a-PbPPO was successfully constructed, and the fusion protein was successfully expressed in Escherichia coli Rosetta. After optimization, it was proved that the fusion protein had the highest expression at 28 ℃ and 1.0 mmol·L^-1 IPTG for 5 h. This study not only created the conditions for the study of PbPPO enzymatic properties, but also provided the scientific basis for the molecular regulation of stone cell content and the improvement of Dangshan Su pear quality.
出处
《亚热带植物科学》
2017年第3期209-214,共6页
Subtropical Plant Science
基金
国家自然科学基金(No.31640068)
关键词
厚砀山酥梨
木质素
石细胞
多酚氧化酶
原核表达
Pyrus bretschneideri ＇Dangshan Su＇
lignin
stone cells
PPO
prokaryotic expression
