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mutation breeding of high 4-androstene-3,17-dione-producing Mycobacterium neoaurum ZADF-4 by atmospheric and room temperature plasma treatment

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2015年第4期16卷 286-295页

作者： Chao LIU Xian ZHANG Zhi-ming RAO Ming-long SHAO Le-le ZHANG Dan WU Zheng-hong XU Hui LI Key Laboratory of Industrial Biotechnology Ministry of Education School of Biotechnology Jiangnan University Laboratory of Applied Microorganisms and Metabolic Engineering School of Biotechnology Jiangnan University Laboratory of Pharmaceutical Engineering School of Pharmaceutical Science Jiangnan University

Steroid medication is used extensively in clinical applications and comprises a large and vital part of the pharmaceutical industry. However, the difficulty of separating 4-androstene-3,17-dione （AD） from 1,4-androstadiene- 3,17-dione （ADD） restricts the application of the microbial transformation of phytosterols in the industry. A novel at- mospheric and room temperature plasma （ARTP） treatment, which employs helium as the working gas, was used to generate Mycobacterium neoaurum mutants producing large amounts of AD. After treatment of cultures with ARTP, four mutants were selected using a novel screening method with a color assay. Among the mutants, M. neoaurum ZADF-4 was considered the best candidate for industrial application. When the fermentation medium contained 15 g/L phytosterols and was cultivated on a rotary shaker at 160 r/min at 30 ~C for 7 d, （6.28＋0.11） g/L of AD and （0.82＋0.05） g/L of ADD were produced by the ZADF-4 mutant, compared with （4.83＋0.13） g/L of AD and （2.34＋0.06） g/L of ADD by the original strain, Iv~. neoaururn ZAD. Compared with ZAD, the molar yield of AD increased from 48.3% to 60.3% in the ZADF-4 mutant. This result indicates that ZADF-4 may have potential for industrial production of AD.

关键词： Mycobacterium neoaurum Atmospheric and room temperature plasma (ARTP) mutation breeding 4-Androstene-3 17-dione (AD) 1 4-Androstadiene-3 17-dione (ADD)

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mutation breeding of Lactobacillus and Its Application in Yoghurt

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Agricultural Biotechnology 2023年第1期12卷 31-34页

作者： Xin LI Shiyun TANG Fengrong TIAN Bing ZHANG Shijun FU Shijin GUO Shandong Binzhou Animal Science&Veterinary Medicine Academy Binzhou 256600China Shandong Anyouxian Food Co. Ltd.Binzhou 256600China Shandong Lvdu Ante Veterinary Drug Industry Binzhou 256600China

With the gradual improvement of the quality of people’s life, the disorder of lipid metabolism caused by high-fat diet has become the main reason that affects the health of our people. Hyperlipidemia, hypertension and hyperglycemia have the effect of causing a large number of serious complications, and are also the main factors of cardiovascular disease. Lactic acid bacteria can avoid hyperlipidemia by inhibiting the excessive growth of harmful intestinal bacteria, maintaining the balance of intestinal microecology, absorbing intestinal cholesterol and regulating lipid metabolism. Therefore, this paper focused on the mutation and breeding of Lactobacillus, and analyzed its application in yoghurt, aiming to provide reference for relevant researchers.

关键词： Lactobacillus mutation breeding Yogurt Application

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A Mutant of Bacillus Subtilis with High-Producing Surfactin by Ion Beam Implantation

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Plasma Science and Technology 2006年第4期8卷 491-496页

作者：刘清梅袁航王军贡国鸿周伟樊永红王丽姚建铭余增亮 Key Laboratory of Ion Beam Bioengineering Chinese Academy of SciencesHefei 230031China

In order to generate a mutant of Bacillus subtilis with enhanced surface activity through low energy nitrogen ion beam implantation, the effects of energy and dose of ions implanted were studied. The morphological changes in the bacteria were observed by scanning electron microscope （SEM）. The optimum condition of ions implantation, 20 keV of energy and 2.6 × 10^15N^＋/cm^2 in dose, was determined. A mutant, B.s-E-8 was obtained, whose surface activity of 50-fold and 100-fold diluted cell-free Landy medium was as 5.6-fold and 17.4-fold as the wild strain. The microbial growth and biosurfactant production of both the mutant and the wild strain were compared. After purified by ultrafiltration and SOURCE 15PHE, the biosurfactant was determined to be a complex of surfactin family through analysis of electrospray ionization mass spectrum （ESI/MS） and there was an interesting finding that after the ion beam implantation the intensities of the components were different from the wild type strain.

关键词： bacillus subtilis surfactin low energy ion beam implantation mutation breeding

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Screening of Bioflocculant-Producing Strain by Ion Implantation and Flocculating Characteristics of Bioflocculants

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Plasma Science and Technology 2008年第3期10卷 394-397页

作者：李培睿李宗伟李宗义秦广雍霍裕平 Key Laboratory of Ion Beam Bio-Engineering Zhengzhou University Zhengzhou 450052 China College of Life Sciences Henan Normal University Xinxiang 453002 China Key Laboratory of Ion Beam Bio-Engineering Zhengzhou University Zhengzhou 450052 China College of Life Sciences Henan Normal University Xinxiang 453002 China

A bioflocculant-producing mutator strain, NIM-192, was screened out through nitrogen ion implanting into F J-7 strain. The results showed that NIM-192 had good genetic stability and high flocculating activity, and the flocculating rate increased by 34.26% than that of the original. Sucrose, complex nitrogen source contained yeast extract, urea and pH 7.0～ 9.0 were chosen as the best carbon source, nitrogen source and initial solution pH for bioflocculant production, respectively. The bioflocculant kept high and stable flocculating activity at alkalinous reaction mixture with a pH beyond 7.0, while the flocculating activity was remarkably reduced when the reaction pH was lower than 7.0. Addition of many cations could obviously increase the flocculating rate, among which Ca^2＋ demonstrated the best effect. The bioflocculant had very strong acid-base stability and *** flocculating rate kept over 86% when pH of the bioflocculant was in a range of 3.0 ～ 12.0, and the change of flocculating activity was not great when heated at 100℃ for 60 rain.

关键词： ion implantation bioflocculant mutation breeding flocculating characteristics

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Morphological and biochemical responses of Oryza sativa L.(cultivar MR219) to ion beam irradiation

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2013年第12期14卷 1132-1143页

作者： Anna Pick Kiong LING Ying Chian UNG Sobri HUSSEIN Abdul Rahim HARUN Atsushi TANAKA Hase YOSHIHIRO Division of Human Biology International Medical University (IMU)57000 Kuala Lumpur Malaysia Department of Science Faculty of Engineering and Science Universiti Tunku Abdul Rahman53300 Setapak Kuala Lumpur Malaysia Agrotechnology and Bioscience Division Malaysian Nuclear Agency Bangi43000 Kajang Selangor Malaysia Radiation-Applied Biology Division Quantum Beam Science Directorate Japan Atomic Energy Agency 1233 Watanuki-machiTakasaki Gunma 370-1292 Japan

Objective: Heavy ion beam, which has emerged as a new mutagen in the mutation breeding of crops and ornamental plants, is expected to result in the induction of novel mutations. This study investigates the morphological and biochemical responses of Oryza sativa toward different doses of carbon ion beam irradiation. Methods: In this study, the dry seeds of O. sativa were irradiated at 0, 20, 40, 60, 80, 100, and 120 Gy, followed by in-vitro germination under controlled conditions. Morphological and biochemical studies were conducted to investigate the morphological and physiological responses of O. sativa towards ion beam irradiation. Results: The study demonstrated that low doses(10 Gy) of ion beam have a stimulating effect on the height, root length, and fresh weight of the plantlets but not on the number of leaves. Meanwhile, doses higher than 10 Gy caused reductions in all the morphological parameters studied as compared to the control samples. The highest total soluble protein content [(2.11±0.47) mg/g FW] was observed in plantlets irradiated at 20 Gy. All irradiated plantlets were found to have 0.85% to 58.32% higher specific activity of peroxidase as compared to the control samples. The present study also revealed that low doses of ion beam(10 and 20 Gy) had negligible effect on the total chlorophyll content of O. sativa plantlets while 40 Gy had a stimulating effect on the chlorophyll content. Plantlets irradiated between 40 to 120 Gy were shown to be 0.38% to 9.98% higher in total soluble nitrogen content which, however, was not significantly different from the control samples. Conclusions: Carbon ion beam irradiation administered at low to moderate doses of 10 to 40 Gy may induce O. sativa mutants with superior characteristics.

关键词： In vitro mutagenesis Ion beam irradiation Total chlorophyll content Total soluble protein content mutation breeding

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Screening of flocculant-producing strains by NTG mutagenesis

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Journal of Environmental Sciences 2005年第3期17卷 494-498页

作者： HUANGXiao-wu CHENGWen HUYong-you SchoolofEnvironmentalScienceandEngineering SouthChinaUniversityofTechmologyGuangzhou510640China SchoolofEnvironmentalScienceandEngineering SouthChinaUniversityofTechmologyGuangzhou510640China

Screening of new microorganism being able to produce efficiently flocculants was carried out. A new model for screening efficient flocculant-producing strains was designed and tested. The results showed that this model for screening efficient flocculant-producing strains is very reliable and can greatly shorten the screening period. 13 flocculant-producing strains were isolated from activated sludge by conventional method. A strain, designated as HHE6, produced the bioflocculant with the turbidity removal 98% for kaolin suspension. Six of 13 strains selected as the original strains were treated with NTG as mutagen, and five mutant strains(HHE-P7, HHE-A8, HHE-P21, HHE-P24, HHE-A26) with high flocculation efficiency was obtained by selection, which exhibited the flocculation rate for kaolin suspension above 90%. Strains HHE6, HHE-P7, and HHE-P24 were classified as Penicillium purpurogenum, HHE-P21 as Penicillium cyclopium, HHE-A26 as Aspergillus versicolor and HHE-A8 as Aspergillus fumigatus, and it is hitherto unreported for biofloccutant-producing strains of Penicillium. The growth of the six strains(HHE6, HHE-P7, HHE-A8, HHE-P21, HHE-P24, HHE-A26) had similar curves, i.e. firstly increasing rapidly, keeping relatively constant then and finally decreasing gradually with cultivation time. The production of bioflocculants by strains showed the similar pattern to strain growth.

关键词： bioflocculant mutation breeding bioflocculant-producing strains Penicillium Aspergillus

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A Comparative Study on Mutagenic Effects of Space Flight and Irradiation of γ-rays on Rice

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Agricultural Sciences in China 2006年第11期5卷 812-819页

作者： WEI Li-jun XU Jian-long WANG Jun-min YANG Qian LUO Rong-ting ZHANG Ming-xian BAO Gen- liang SUN Ye-qing Department of Life Science and Engineering College of Science Harbin Institute of Technology Harbin 150001 P.R.China The Institute of Crop Sciences/The National Key Facility for Crop Gene Resources and Genetic Improvement Chinese Academy of Agricultural Sciences Beijing 100081 P.R.China Institute of Crop and Nuclear Technology Utilization Zhejiang Academy of Agricultural Sciences Hangzhou 310021 P.R.China

The experiment was conducted to study the mutagenic effects of space environment on seedling growth in M1 generation and plant height and heading date in M2 generation in rice. Two types of lines of growth promotion （GP） and growth suppression （GS） were selected from the recombinant inbred lines （RILs） derived from a cross between a sensitive japonica Lemont and a nonsensitive indica Teqing to compare the mutation frequency and mutagenic efficiency for plant height and heading date in the M2 generation. Space environment resulted in 34.9% higher seedling height （SH） in the GP group than in the control, and 39.1% lower in the GS group than in the control, and there was no difference in seed fertility （SF） between the two groups. In M2, mutants of plant height and heading date can be induced by space treatment in both the two groups with lower mutation frequency and mutagenic efficiency in the GP group than in the GS group. There were no significant differences in the physiological damages in M1 between the two groups after γ-rays irradiation treatment. mutation frequency and mutagenic efficiency of heading date in M2 were higher in the GS group than in the GP group, and the opposite was true for mutagenic efficiency of plant height although the mutation frequency varied between the two groups. The mutation frequencies of plant height and heading date induced by space environment were obviously lower than those by γ-rays irradiation, but the mutagenic efficiency was the opposite for most of the traits. For the GP and GS of seedlings induced by space environment, the GS had higher mutation frequency and mutagenic efficiency for plant height and heading date than the GP in M2.

关键词： rice space environment-induced mutation mutagenic effect physiological damage mutation breeding

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Frequency and type of inheritable mutations induced by γ rays in rice as revealed by whole genome sequencing

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Journal of Zhejiang University-Science B(Biomedicine & Biotechnology) 2016年第12期17卷 905-915页

作者： Shan LI Yun-chao ZHENG Hai-rui CUI Hao-wei FU Qing-yao SHU Jian-zhong HUANG National Key Laboratory of Rice Biology Institute of Crop Sciences Zhejiang University Hangzhou 310058 China Hubei Collaborative Innovation Center for Grain Industry Jingzhou 434025 China Institute of Nuclear Agricultural Sciences Zhejiang University Hangzhou 310029 China Jiaxing Academy of Agricultural Sciences Jiaxing 314016 China

mutation breeding is based on the induction of genetic variations; hence knowledge of the frequency and type of induced mutations is of paramount importance for the design and implementation of a mutation breeding program. Although γ ray irradiation has been widely used since the 1960 s in the breeding of about 200 economically important plant species, molecular elucidation of its genetic effects has so far been achieved largely by analysis of target genes or genomic regions. In the present study, the whole genomes of six γ-irradiated M2 rice plants were sequenced; a total of 144–188 million high-quality（Q〉20） reads were generated for each M2 plant, resulting in genome coverage of 45 times for each plant. Single base substitution（SBS） and short insertion/deletion（Indel） mutations were detected at the average frequency of 7.5×10^-6～9.8×10^-6 in the six M2 rice plants（SBS being about 4 times more frequent than Indels）. Structural and copy number variations, though less frequent than SBS and Indel, were also identified and validated. The mutations were scattered in all genomic regions across 12 rice chromosomes without apparent hotspots. The present study is the first genome-wide single-nucleotide resolution study on the feature and frequency of γ irradiation-induced mutations in a seed propagated crop; the findings are of practical importance for mutation breeding of rice and other crop species.

关键词： mutation breeding γ rays mutation spectrum Genomic variation

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breeding of High-Yield Salinomycin-Producing Streptomyces albus Strains by Low Energy N^+ Ion Beam Irradiation

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Agricultural Biotechnology 2012年第5期1卷 55-56,F0003页

作者： Liqin WU Fengxiang MIAO Haike GU Hongzhong SHANG Beijing Radiation Center Heze Lukang Shelile Pharmaceutical Co. Ltd

[Objective] This study aimed to explore the mutagenesis effects of N＋ ion beam implantation on Streptomyces a/bus and obtain high-yield salinomycin- producing mutant strain. [ Method ] Streptomyces a/bus strain S-11-04 was mutated with different doses of N ＋ implantation. The effects of low energy N ＊ implantation on the survival rate, colony morphology and salinomycin-producing ability were investigated. [ Result] The results showed that low energy N ＋ implantation can efficiently improve the positive mutation rate of Streptomyces albus; 13 mutant strains with high yield of salinomycin were isolated; to be specific, mutant strain N3- 6 has relatively good genetic stability with four continuous generations, and the titres of salinomycin were increased by 41% in the shake-flask culture and 20.5% in mass production compared with the control. [ Conclusion ] N ＋ ion beam irradiation is an effective method to obtain high-yield salinomycin-producing Streptomy- ces albus strain.

关键词： Streptomyces albus N + ion beam irradiation Salinomycin mutation breeding

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Enhanced production of L-cysteine by mutant Pseudomonas sp.C-25 and structural prediction of its L-NCC hydrolase

Enhanced production of L-cysteine by mutant Pseudomonas sp.C...

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2015中国酶工程与糖生物工程学术研讨会

作者： Xu Chen Weiqing Chen Jin Huang Pu Wang College of Pharmaceutical Science Zhejiang University of Technology College of Biological and Environmental Engineering Zhejiang Shuren University

Pseudomonas sp.B-3 is a promising industrial strain for bioconversion of DL-2-amino-△-thiazoline-4-carboxylic acid(DZ-ATC)to *** order to further enhance its bioconversion ability,nitrogen ion beam implantation was applied in strain B-3 and a powerful mutant with highest enzyme activity designated as Pseudomonas sp.C-25 was obtained,of which Z-cysteine enzyme activity achieved 2748 U/mL,an increase of 26.3%compared to that of initial strain B-3(2176 U/mL).Meanwhile the yield of Z-cysteine improved from 11.85 g/L to 14.12 g/*** mutant strain exhibited favorable property about producing Z-cysteine over the original *** maximum enzyme activity of strain C-25 in 7 L fermentor reached 3925 U/mL,and the cultivation time with maximum enzyme activity in the fermentor was shorten for 11 h than in the Erlenmeyer flasks,and the yield of Z-cysteine attained 16.53 g/L in 7 L ***,the genes of N-carbamyl-Z-cysteine(Z-NCC)hydrolase were cloned from strain B-3 and mutant strain C-25 respectively and their protein sequence were ***,online server I-TASSER was used to predict its 3-dimensional(3D)structure and understand the function *** results showed that the first mutant site was 212nucleotide,which changed from T to C and the 71amino acid changed from Ile to Thr,the second mutant site was 1052nucleotide,which changed from C to T and the 351amino acid changed from His to *** protein 3D structure of Z-NCC hydrolase was predicted and has a high confidence.

关键词： L-cysteine Pseudomonas sp. N+ ion beam implantation mutation breeding N-carbamyl-L-cysteine(L-NCC) hydrolase modeling

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