产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte
为了探究细菌异质性乙酰辅酶A羧化酶β亚基生物活性及其对乙酰辅酶A羧化酶酶活影响，以高产油核桃内生细菌B. subtilis HB1310基因组DNA为模板，利用PCR技术扩增其乙酰辅酶A羧化酶羧基转移酶β亚基（β-CT）基因（acb基因），构建pET-28a-acb载体并在E. coli BL21中表达，进一步对乙酰辅酶A羧化酶acb基因的诱导表达条件进行了优化。结果表明：乙酰辅酶A羧化酶acb基因在E. coli BL21中实现了表达，分子质量在25~35 kDa之间；重组蛋白最佳诱导表达条件为诱导剂异丙基-β-D-硫代半乳糖苷（IPTG）浓度1.0 mmol/L、诱导时间6 h、诱导初始p...
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中粮工科（西安）国际工程有限公司
2023-04-01
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| Series: | Zhongguo youzhi |
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| Online Access: | http://tg.chinaoils.cn/ch/reader/create_pdf.aspx?file_no=20230418&year_id=2023&quarter_id=4&falg=1 |
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| author | 许思远，张永贵，赵沛，曹娟娟，张琴XU Siyuan，ZHANG Yonggui，ZHAO Pei，CAO Juanjuan，ZHANG Qin |
| author_facet | 许思远，张永贵，赵沛，曹娟娟，张琴XU Siyuan，ZHANG Yonggui，ZHAO Pei，CAO Juanjuan，ZHANG Qin |
| author_sort | 许思远，张永贵，赵沛，曹娟娟，张琴XU Siyuan，ZHANG Yonggui，ZHAO Pei，CAO Juanjuan，ZHANG Qin |
| collection | DOAJ |
| description | 为了探究细菌异质性乙酰辅酶A羧化酶β亚基生物活性及其对乙酰辅酶A羧化酶酶活影响，以高产油核桃内生细菌B. subtilis HB1310基因组DNA为模板，利用PCR技术扩增其乙酰辅酶A羧化酶羧基转移酶β亚基（β-CT）基因（acb基因），构建pET-28a-acb载体并在E. coli BL21中表达，进一步对乙酰辅酶A羧化酶acb基因的诱导表达条件进行了优化。结果表明：乙酰辅酶A羧化酶acb基因在E. coli BL21中实现了表达，分子质量在25~35 kDa之间；重组蛋白最佳诱导表达条件为诱导剂异丙基-β-D-硫代半乳糖苷（IPTG）浓度1.0 mmol/L、诱导时间6 h、诱导初始pH 8.0，在此条件下工程菌的乙酰辅酶A羧化酶活性可达（4.11±0.03） U/mL，较野生菌提高39.7%。综上，乙酰辅酶A羧化酶β-CT为具有较好生物活性的乙酰辅酶A羧化酶亚基。
To explore the biological activity of the bacterial heterogeneous acetyl-CoA carboxylase β subunit and its influence on the acetyl-CoA carboxylase activity, the acetyl-CoA carboxylase carboxyltransferase subunit beta(β-CT) gene(acb gene) was amplified by PCR technology using the genomic DNA of a high oil-producing walnut endophyte B. subtilis HB1310 as template, the expression vector pET-28a-acb was constructed and expressed in E. coli BL21, and the induced expression conditions of this acb gene were further optimized. The results showed that the acetyl-CoA carboxylase acb gene was expressed in E. coli BL21 with a molecular weight of 25-35 kDa. Moreover, the optimal induced expression conditions were determined as follows: isopropyl-beta-D-thiogalactopyranoside (IPTG) concentration 1.0 mmol/L, induction time 6 h, and induction initial pH 8.0. Under the optimal conditions, the acetyl-CoA carboxylase activity of the engineering bacteria reached (4.11±0.03) U/mL, which was 39.7% higher than that of the wild bacteria. In conclusion, acetyl-CoA carboxylase β-CT is an acetyl-CoA carboxylase subunit with good biological activity. |
| first_indexed | 2024-03-13T02:36:20Z |
| format | Article |
| id | doaj.art-6fcf7a13c5d84712b8e2a3ccf419af27 |
| institution | Directory Open Access Journal |
| issn | 1003-7969 |
| language | English |
| last_indexed | 2024-03-13T02:36:20Z |
| publishDate | 2023-04-01 |
| publisher | 中粮工科（西安）国际工程有限公司 |
| record_format | Article |
| series | Zhongguo youzhi |
| spelling | doaj.art-6fcf7a13c5d84712b8e2a3ccf419af272023-06-29T01:22:29Zeng中粮工科（西安）国际工程有限公司Zhongguo youzhi1003-79692023-04-0148499104https://doi.org/10.19902/j.cnki.zgyz.1003-7969.220108产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte许思远，张永贵，赵沛，曹娟娟，张琴XU Siyuan，ZHANG Yonggui，ZHAO Pei，CAO Juanjuan，ZHANG Qin0安徽工程大学 生物与食品工程学院，安徽 芜湖 241000College of Biological and Food Engineering, Anhui Polytechnic University, Wuhu 241000, Anhui, China为了探究细菌异质性乙酰辅酶A羧化酶β亚基生物活性及其对乙酰辅酶A羧化酶酶活影响，以高产油核桃内生细菌B. subtilis HB1310基因组DNA为模板，利用PCR技术扩增其乙酰辅酶A羧化酶羧基转移酶β亚基（β-CT）基因（acb基因），构建pET-28a-acb载体并在E. coli BL21中表达，进一步对乙酰辅酶A羧化酶acb基因的诱导表达条件进行了优化。结果表明：乙酰辅酶A羧化酶acb基因在E. coli BL21中实现了表达，分子质量在25~35 kDa之间；重组蛋白最佳诱导表达条件为诱导剂异丙基-β-D-硫代半乳糖苷（IPTG）浓度1.0 mmol/L、诱导时间6 h、诱导初始pH 8.0，在此条件下工程菌的乙酰辅酶A羧化酶活性可达（4.11±0.03） U/mL，较野生菌提高39.7%。综上，乙酰辅酶A羧化酶β-CT为具有较好生物活性的乙酰辅酶A羧化酶亚基。 To explore the biological activity of the bacterial heterogeneous acetyl-CoA carboxylase β subunit and its influence on the acetyl-CoA carboxylase activity, the acetyl-CoA carboxylase carboxyltransferase subunit beta(β-CT) gene(acb gene) was amplified by PCR technology using the genomic DNA of a high oil-producing walnut endophyte B. subtilis HB1310 as template, the expression vector pET-28a-acb was constructed and expressed in E. coli BL21, and the induced expression conditions of this acb gene were further optimized. The results showed that the acetyl-CoA carboxylase acb gene was expressed in E. coli BL21 with a molecular weight of 25-35 kDa. Moreover, the optimal induced expression conditions were determined as follows: isopropyl-beta-D-thiogalactopyranoside (IPTG) concentration 1.0 mmol/L, induction time 6 h, and induction initial pH 8.0. Under the optimal conditions, the acetyl-CoA carboxylase activity of the engineering bacteria reached (4.11±0.03) U/mL, which was 39.7% higher than that of the wild bacteria. In conclusion, acetyl-CoA carboxylase β-CT is an acetyl-CoA carboxylase subunit with good biological activity.http://tg.chinaoils.cn/ch/reader/create_pdf.aspx?file_no=20230418&year_id=2023&quarter_id=4&falg=1核桃内生细菌；产油；乙酰辅酶a羧化酶；克隆；优化表达walnut endophyte; oil-producing; acetyl-coa carboxylase; cloning; expression optimization |
| spellingShingle | 许思远，张永贵，赵沛，曹娟娟，张琴XU Siyuan，ZHANG Yonggui，ZHAO Pei，CAO Juanjuan，ZHANG Qin 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte Zhongguo youzhi 核桃内生细菌；产油；乙酰辅酶a羧化酶；克隆；优化表达 walnut endophyte; oil-producing; acetyl-coa carboxylase; cloning; expression optimization |
| title | 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte |
| title_full | 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte |
| title_fullStr | 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte |
| title_full_unstemmed | 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte |
| title_short | 产油核桃内生细菌乙酰辅酶A羧化酶acb基因 克隆及优化表达Cloning and expression optimization of acetyl-CoA carboxylase acb gene from oil-producing walnut endophyte |
| title_sort | 产油核桃内生细菌乙酰辅酶a羧化酶acb基因 克隆及优化表达cloning and expression optimization of acetyl coa carboxylase acb gene from oil producing walnut endophyte |
| topic | 核桃内生细菌；产油；乙酰辅酶a羧化酶；克隆；优化表达 walnut endophyte; oil-producing; acetyl-coa carboxylase; cloning; expression optimization |
| url | http://tg.chinaoils.cn/ch/reader/create_pdf.aspx?file_no=20230418&year_id=2023&quarter_id=4&falg=1 |
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