桑树查尔酮异构酶基因的克隆与原核表达分析

Liu Changying, Southwest University, Chongqing,College of Biotechnology; Zhao Aichun, Southwest University, Chongqing,College of Biotechnology; Li Jun, Southwest University, Chongqing,College of Biotechnology

Cloning and Prokaryotic Expression of Chalcone Isomerase Gene from Mulberry （Morus alba） | 桑树查尔酮异构酶基因的克隆与原核表达分析

2013

Liu Changying, Southwest University, Chongqing(China),College of Biotechnology | Zhao Aichun, Southwest University, Chongqing(China),College of Biotechnology | Li Jun, Southwest University, Chongqing(China),College of Biotechnology

中国人. 以果叶兼用新桑品种‘嘉陵30号’的果实为材料,采用同源克隆法和抑制PCR法克隆桑树查尔酮异构酶基因（MaCHI）全序列。该基因的基因组序列全长2402bp,包含2112bp长的ORF和290bp长的3＇UTR序列,ORF由4个外显子和3个内含子组成,该基因编码219个氨基酸。预测MaCHI编码蛋白质的分子质量为23.8ku,理论等电点为5.29。采用RT-PCR法分析MaCHI在不同组织中的表达水平,在叶片和果中的表达量较高,在根中表达量较低。构建pET-28a（＋）-MaCHI原核表达重组质粒,并在大肠杆菌中诱导表达。

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英语. The MaCHI gene was cloned from the ripe fruit of Morus alba ‘Jialing 30’ by homologous cloning and suppression PCR. The full-length genomic sequence of MaCHI is 2 402 bp in length, with the 3′UTR of 290 bp and the open reading frame （ORF） of 2 112 bp. Its ORF consists of four exons and three introns, and its cDNA encodes 219 amino acids. The putative molecular weight of the protein encoded by MaCHI gene is 23.8 ku and its theoretical isoelectric point is 5.29. The RT-PCR was used to analyze the expression levels of MaCHI in different tissues, and its expression level was high in leaves and fruits, and relatively low in roots. A prokaryotic expression recombinant plasmid, pET-28a （＋）-MaCHI was constructed and transformed into Escherichia coli for expression. This study will help establish theoretical foundation for the further research of MaCHI.

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