Cloning, Spatiotemporal Expression and Prokaryotic Expression of ACC Oxidase Gene (BnACO2) in Ramie
1 Ramie Research Institute, Hunan Agricultural University, Changsha 410128, China
2 Hunan Provincial Key Laboratory of Germplasm Resource Innovation and Resource Utilization, Changsha, 410128, China
Plant Gene and Trait, 2021, Vol. 12, No. 1
Received: 03 Jan., 2021 Accepted: 15 Jan., 2021 Published: 22 Jan., 2021
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This article was first published in Molecular Plant Breeding in Chinese, and here was authorized to translate and publish the paper in English under the terms of Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
ACC oxidase (ACO) is a key enzyme in ethylene synthesis pathway. This study by rt-pcr with RACE technology get a ACO gene cloning, named BnACO2, total length of the gene cDNA sequence is 1 377 bp, the open reading frame of 957 bp, coded 318 amino acid polypeptide, predict its molecular weight and isoelectric point (pl) 36.145 kD and 5.60, respectively, bioinformatics analysis showed that BnACO2 no encoding protein signal peptide and transmembrane domain structure, subcellular localization in the cytoplasm. The similarity of ACO gene nucleotide sequence and amino acid sequence was more than 82% and 84% respectively. Phylogenetic analysis showed that the ACO gene was closely related to jute and hemp. Real-time fluorescence quantitative PCR analysis showed that BnACO2 gene was expressed in all parts of ramie, especially in female flower buds. The prokaryotic expression vector PQe-BnACO2 was successfully constructed by double enzyme digestion. At 37°C, 1.0mmol/L IPTG induced PQe-BnACO2 to express the target protein, and the molecular weight of the induced protein was about 36.15kda, which was consistent with the predicted protein size. This lays the foundation for further study on the function of BnACO2 gene.
Ramie; ACC oxidase; Clone; Expression
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