[1]丁 强,向继武,周 明,等.气液相等离子体诱变选育安普霉素高产菌株的研究[J].武汉工程大学学报,2023,45(05):510-516.[doi:10.19843/j.cnki.CN42-1779/TQ.202307018]
　DING Qiang,XIANG Jiwu,ZHOU Ming,et al.Gas-Liquid Phase Plasma Mutagenesis Breeding and High-Yield Apramycin Mutant Strain Screening[J].Journal of Wuhan Institute of Technology,2023,45(05):510-516.[doi:10.19843/j.cnki.CN42-1779/TQ.202307018]

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气液相等离子体诱变选育安普霉素高产

菌株的研究

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《武汉工程大学学报》[ISSN:1674-2869/CN:42-1779/TQ]

卷:

45

期数:

2023年05期

页码:

510-516

栏目:

化学与化学工程

出版日期:

2023-11-17

文章信息/Info

Title:

Gas-Liquid Phase Plasma Mutagenesis Breeding and High-Yield Apramycin Mutant Strain Screening

文章编号:

1674 - 2869（2023）05 - 0510 - 07

作者:

丁 强¹; 向继武¹; 周 明¹; 程 波²; 谭小芳¹ ; 闫 雪^*3;  沈 洁⁴

1. 宜昌三峡制药有限公司，湖北 宜昌 443000；

2. 武汉工程大学环境生态与生物工程学院，湖北 武汉 430205；

3. 宜昌人福药业有限责任公司，湖北 宜昌 443000 ；

4. 中国科学院合肥物质科学研究院，安徽 合肥 230000

Author(s):

DING Qiang¹; XIANG Jiwu¹; ZHOU Ming¹; CHENG Bo²; TAN Xiaofang¹; YAN Xue^*3; SHEN Jie⁴

1. Yichang Sanxia Pharmaceutical Co.， Ltd， Yichang 443000，China；

2.School of Environmental Ecology and Biological Engineering， Wuhan Institute of Technology，Wuhan 430205，China；

3. Yichang Humanwell Pharmaceutical Co.， Ltd，Yichang 443000，China；

4. Hefei Institutes of Physical Science， Chinese Academy of Sciences，Hefei 230000，China

关键词:

安普霉素; 黑暗链霉菌; 气液相等离子体诱变; 筛选

Keywords:

apramycin;  streptomyces tenebrarius;  gas-liquid phase plasma mutagenesis;  screening

分类号:

Q812

DOI:

10.19843/j.cnki.CN42-1779/TQ.202307018

文献标志码:

A

摘要:

以宜昌三峡制药有限公司保藏的黑暗链霉菌（Streptomyces tenebrarius）菌株A-0为出发菌种，采用改进后的气液相等离子体诱变法，选育安普霉素稳定高产突变菌株。诱变后挑取单孢子获得生长平板后，采用24孔板快速筛选法初筛后摇瓶复筛，高效液相色谱法进行安普霉素含量检测及成品质量分析，同时结合杯碟法（枯草芽孢杆菌（bacillus subtitles）为指示菌）进行生物效价测定。结果显示：气液相等离子体诱变对安普霉素菌株突变效果较显著，处理75 s后，致死率为78.6%时正突变率达28.9%。孔板初筛得到5株效价增幅5%~30%的突变株，复筛后获得1株安普霉素稳定突变株A-4，其发酵水平达到0.375 mg/mL，生物效价达到3 420 U/mL，较出发菌株生物效价提高了32.6%。经5代传代，发酵水平变化低于5%。结果表明：诱变用的活性离子导致细胞DNA损伤和破坏，在其自我修复的过程中形成了不完全修复的突变，最终获得了稳定遗传的突变菌株。

Abstract:

The original strain Streptomyces tenebrarius A-0， preserved by Yichang Sanxia Pharmaceutical Co.， Ltd， was subjected to mutation using an improved gas-liquid phase plasma mutagenesis method. The objective was to obtain a mutant strain of Streptomyces tenebrarius A-0 with a high yield of apramycin. After mutation， single spores were picked and cultured on agar plates. A preliminary screening was conducted with a 24-well plate， followed by a secondary screening via shake flask fermentation. The assay of apramycin yield and analysis of finished sample quality were conducted using a high performance liquid chromatography. Additionally， the bioassay test was performed using the cylinder plate method with Bacillus subtilis as the test organism. The results indicated that gas-liquid phase plasma mutagenesis has a positive impact on the apramycin strain， with a mutation rate of 28.9% observed when the lethality rate is 78.6% after 75 s of treatment. Five mutant strains with 5%-30% increase in bioassay activity were identified during the preliminary screening. Following the secondary screening， a stable and high-yield apramycin mutant strain named A-4 was obtained， exhibiting a fermentation level of 0.375 mg/mL and a bioassay activity of 3 420 U/mL. This represents a 32.6% increase in bioassay activity compared to the original strain. After 5 generations of passage， the fermentation level of the strain A-4 remains stable with less than 5% change. These findings suggested that the active ions utilized in the mutagenesis process induce DNA damage and cell disruption， resulting in incomplete repair mutations during the self-repair mechanism. Finally， a genetically stable mutant strain was obtained.

备注/Memo

备注/Memo:

收稿日期：2023-07-18

作者简介：丁 强，硕士，高级工程师。E-mail：[email protected]

*通讯作者：闫 雪，硕士，工程师。E-mail：[email protected]

引文格式：丁强，向继武，周明，等. 气液相等离子体等离子体诱变选育安普霉素高产菌株的研究［J］. 武汉工程大学学报，2023，45（5）：510-516.

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