JOURNAL OF PLANT PHYSIOLOGY
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2025
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IF 4.1
Yijia Zhong,Zhie Zhou,Zhongping Yin,Lu Zhang,Qingfeng Zhang,Yihai Xie,Jiguang Chen
Physalis peruviana L. ( P. peruviana ) is an edible medicinal plant rich in bioactive phenolics. This study aimed to establish a hairy root (HR) culture of P. peruviana as a potential source for the synthesis of natural compounds. HRs were induced in P. peruviana using different Agrobacterium rhizogenes strains (R1601, C58C1, A4, and K599). Notably, K599 did not induce HR formation, whereas R1601, C58C1, and A4 yielded transformation frequencies of 57.78, 65.14, and 72.31%, respectively. Secondary metabolite production and antioxidant capacity were further examined in HRs induced using C58C1, R1601, and A4. It was found that A. rhizogenes R1601 induced the greatest increase (44% compared to that observed in the non-transformed culture). The methanolic extract of HRs induced by A. rhizogenes R1601 exhibited strong antioxidant capacity, with IC 50 values of 1.41 mg DE/mL and 2.33 mg DE/mL for 2,2-diphenyl-1-picrylhydrazyl and 2,2-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid), respectively. The HR culture showed higher production of phenolic compounds and higher antioxidant capacity than the non-transformed cultures. Ultra-performance liquid chromatography time-of-flight tandem mass spectrometry was used to identify eight alkaloids, phenolics, and glycoside compounds. A. rhizogenes R1601 is emerging as a possible strain for the mass production of HR and bioactive phenolic compounds in P. peruviana .
研究领域:
植物学
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分子生物学
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药用植物研究
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DOI
PubMed
高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2019
·
IF 3.571
Ruiqi Bao,Ning Gao,Jing Lv,Chaofan Ji,Huipeng Liang,Shengjie Li,Chenxu Yu,Zhenyu Wang,Xinping Lin
Nine transformants of Rhodosporidium toruloides with significant changes in the carotenoid profile were obtained by Agrobacterium tumefaciens-mediated transformation, including a white, three red, and four yellow mutants. A red mutant A1-15-BRQ that showed a high torularhodin production was selected for culture condition optimization. Results indicated that the torularhodin yield was boosted with glucose as the carbon source, at a carbon/nitrogen ratio of 22, a loading volume of 75 mL, and 28 °C. The torularhodin yield of 21.3 mg/L consisting of 94.4% total carotenoids was obtained by Box–Behnken design experiments. The torularhodin yield was 17.0-fold higher than that of the wild type, with time shortened from 9 to 3 days. This study reports an effective strategy for improving torularhodin production and provides a candidate R. toruloides strain for highly selective production of torularhodin.
研究领域:
微生物学
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生物技术
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代谢工程
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DOI
PubMed
高影响因子
Plant Methods
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2018
·Vol. 14
·期 1
·1-8
·
IF 4.269
Alagarsamy Karthikeyan,Shamala Lubobi Ferdinand,Wei Shu
Background Camellia sinensis var. sinensis is widely grown for tea beverages that possess significant health promoting effects. Studies on tea plant genetics and breeding are hindered due to its recalcitrance to Agrobacterium -mediated genetic transformation. Among the possible reasons, oxidation of phenolics released from explant tissues and bactericidal effects of tea polyphenols during the process of transformation play a role in the plant recalcitrance. The aim of the present study was to alleviate the harmful effects of phenolic compounds using in - planta transformation. Results Two-month old seedlings of tea cultivar “Nong Kangzao” were infected at the hypocotyl with wild type Agrobacterium rhizogenes and maintained in an environment of high humidity. 88.3% of infected plants developed hairy roots at the wounded site after 2 months of infection. Our data indicated that transgenic hairy root induction of tea can be achieved using A. rhizogenes following the optimized protocol. Conclusion With this method, composite tea plants containing wild-type shoots with transgenic roots can be generated for “in root” gene functional characterization and root-shoot interaction studies. Moreover, this method can be applied to improve the root system of composite tea plants for a better resistance to abiotic and biotic stresses.
研究领域:
植物生物技术
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分子遗传学
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农学
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DOI
PubMed
高影响因子
BIOTECHNOLOGY AND APPLIED BIOCHEMISTRY
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2021
·Vol. 69
·期 4
·1428-1437
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IF 2.431
Di Liu,Linbo Gou,Yajun Bai,Tai-Ping Fan,Xiaohui Zheng,Yujie Cai
In this study, the 3-quinuclidinone reductase from Agrobacterium tumefaciens (AtQR) was modified by site-directed mutagenesis. And we further obtained a saturation mutant library in which the residue 197 was mutated. A single-point mutation converted the wild enzyme that originally had no catalytic activity in reduction of ethyl 4-chloroacetoacetate (COBE) into an enzyme with catalytic activity. The results of enzyme activity assays showed that the seven variants could asymmetrically reduce COBE to ethyl ( S )-4-chloro-3-hydroxybutyrate (( S )-CHBE) with NADH as coenzyme. In the library, the variant E197N showed higher catalytic efficiency than others. The E197N was optimally active at pH 6.0 and 40°C, and the catalytic efficiency ( k cat / K m ) for COBE was 51.36 s –1 ·mM –1 . This study showed that the substrate specificity of AtQR could be changed through site-directed mutagenesis at the residue 197.
研究领域:
生物化学
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酶学
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生物催化
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DOI
PubMed
高影响因子
Plants-Basel
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2025
·Vol. 14
·期 17
·2664
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IF 4.1
Dan Wang,Yin Liu,Yao Zhang,Xiumei Huang,Jiaxuan Wang,Yi Wang,Yue Liu,Chao Yan,Bingsheng Lv,Yue Jia
Welsh onion ( Allium fistulosum L.), a globally significant vegetable, flavoring agent, and phytomedicine resource, has remained unavailable with established transient expression platforms for functional genomic investigations. To address this critical methodological limitation, we present systematically optimized protocols for both Agrobacterium -mediated hairy root transformation and protoplast transient expression systems, achieving significant advances in transformation efficiency for this species. Through systematic optimization of key parameters, including Agrobacterium rhizogenes ( A. rhizogenes ) strain selection (with Ar.Qual demonstrating superior performance), explant type efficacy, bacterial suspension optical density (OD 600 = 0.3), and acetosyringone induction concentration (100 μM), we established a highly efficient stem disc infection methodology, achieving 88.75% hairy root induction efficiency. Subsequent optimization of protoplast isolation protocols identified the optimal enzymatic digestion conditions: 6-h dark digestion of young leaves using 1.0% ( w / v ) Cellulase R-10, 0.7% ( w / v ) Macerozyme R-10, and 0.4 M mannitol, yielding 3.3 × 10 6 viable protoplasts g −1 FW with 90% viability. System functionality validation through PEG-mediated transient transformation demonstrated successful green fluorescent protein (GFP) reporter gene expression, confirmed by fluorescence microscopy. As the first documented transient expression platforms for Welsh onion, these protocols enable essential molecular investigations, including in planta promoter activity profiling, subcellular protein localization, and CRISPR-based genome-editing validation. This methodological breakthrough overcomes previous technical constraints in Welsh onion molecular biology, providing critical tools for accelerated gene functional characterization in this agriculturally important species.
研究领域:
植物生物技术
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分子生物学
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遗传转化
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功能基因组学
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DOI
PubMed
高影响因子
Journal of Fungi
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2022
·Vol. 8
·期 2
·119
·
IF 5.724
Larissa Balabanova,Aleksandra Seitkalieva,Yulia Yugay,Tatiana Rusapetova,Lubov Slepchenko,Anna Podvolotskaya,Margarita Yatsunskaya,Elena Vasyutkina,Oksana Son,Liudmila Tekutyeva,Yury Shkryl
An efficientAgrobacterium-mediated genetic transformation based on the plant binary vector pPZP-RCS2 was carried out for the multiple heterologous protein production in filamentous fungusThermothelomycesthermophilusF-859 (formerlyMyceliophthorathermophilaF-859). The engineered fungusTh.thermophiluswas able to produce plant storage proteins ofZeamays(α-zein Z19) andAmaranthushypochondriacus(albumin A1) to enrich fungal biomass by valuable nutritional proteins and improved amino acid content. The mRNA levels ofz19anda1genes were significantly dependent on their driving promoters: the promoter of tryptophan synthase (PtrpC) was more efficient to expressa1, while the promoter of translation elongation factor (Ptef) provided much higher levels ofz19transcript abundance. In general, the total recombinant proteins and amino acid contents were higher in the Ptef-containing clones. This work describes a new strategy to improve mycoprotein nutritive value by overexpression of plant storage proteins.Keywords:filamentous fungi;Myceliophthorathermophila;mycoprotein;recombinant protein;maize alpha-zein;amaranth albumin A1
研究领域:
微生物学
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分子生物学
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生物技术
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食品科学
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DOI
PubMed
高影响因子
JOURNAL OF BIOTECHNOLOGY
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2021
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IF 3.307
Yuanyuan Miao,Yanyun Hu,Shanyong Yi,Xuejia Zhang,Ninghua Tan
Rubia yunnanensis is an important medicinal plant with various bioactive secondary metabolites. In order to reduce the dependence on wild populations of the species, we aim to establish in vitro culture system that can produce Rubiaceae-type cyclopeptides (RAs) and quinones. Agrobacterium rhizogenes -mediated transformation of stem segments of in vitro grown R. yunnanensis plants using four A. rhizogenes strains was studied and transformation conditions were optimized. Hairy roots appeared with the highest frequency (68.89%) when stem segments (with leaves) without pre-culture were immersed in A. rhizogenes A4 strain bacterial suspension for 30 min, co-cultured on Murashige and Skoog (MS) solid medium in the dark for three days, and afterwards incubated in darkness. PCR analysis of rol B and rol C genes confirmed transformed nature of six hairy root clones. The hairy roots grew rapidly, especially showing the highest accumulation of biomass in MS liquid medium compared to in vitro grown plants and calli. Histological observation of hairy root revealed anatomical difference in vascular cylinder, where the cells exhibited high mitotic activity characterized by vigorous growth. The UPLC-MS/MS analysis revealed that the amount of RAs in the hairy roots grown in ½MS liquid medium (4.611 μg g −1 DW) was higher than that in in vitro grown plants (0.331 and 4.096 μg g −1 DW for shoots and roots respectively) and calli (1.082 μg g −1 DW), but still far lower than that in the roots of seed-borne plants (80.296 μg g −1 DW). However, the hairy roots accumulated high level of quinones (2320.923 and 5067.801 μg g −1 DW for MS and ½MS liquid media respectively), of the same order of magnitude as the roots of seed-borne plants (7409.973 μg g −1 DW). Hairy root culture of R. yunnanensis , with high accumulation of biomass and production of quinones, may offer an attractive perspective for the production of the RAs and quinones that could be further optimized for pharmaceutical use.
研究领域:
植物生物技术
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生药学
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分子生物学
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药物化学
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产品关联
DOI
PubMed
高影响因子
APPLIED AND ENVIRONMENTAL MICROBIOLOGY
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2019
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IF 4.077
Rongshui Wang,Jihong Yi,Jinmeng Shang,Wenjun Yu,Zhifeng Li,Haiyan Huang,Huijun Xie,Shuning Wang
Agrobacterium tumefaciens S33 degrades nicotine via a novel hybrid of the pyridine and the pyrrolidine pathways. The hybrid pathway consists of at least six steps involved in oxidoreductive reactions before the N-heterocycle can be broken down. Collectively, the six steps allow electron transfer from nicotine and its intermediates to the final acceptor O2 via the electron transport chain (ETC). 6-Hydroxypseudooxynicotine oxidase, renamed 6-hydroxypseudooxynicotine dehydrogenase in this study, has been characterized as catalyzing the fourth step using the artificial electron acceptor 2,6-dichlorophenolindophenol. Here, we used biochemical, genetic, and liquid chromatography-mass spectrometry (LC-MS) analyses to determine that 6-hydroxypseudooxynicotine dehydrogenase utilizes the electron transfer flavoprotein (EtfAB) as the physiological electron acceptor to catalyze the dehydrogenation of pseudooxynicotine, an analogue of the true substrate 6-hydroxypseudooxynicotine, in vivo, into 3-succinoyl-semialdehyde-pyridine. NAD(P)+, O2, and ferredoxin could not function as electron acceptors. The oxygen atom in the aldehyde group of the product 3-succinoyl-semialdehyde-pyridine was verified to be derived from H2O. Disruption of the etfAB genes in the nicotine-degrading gene cluster decreased the growth rate of A. tumefaciens S33 on nicotine but not on 6-hydroxy-3-succinoylpyridine, an intermediate downstream of the hybrid pathway, indicating the requirement of EtfAB for efficient nicotine degradation. The electrons were found to be further transferred from the reduced EtfAB to coenzyme Q by the catalysis of electron transfer flavoprotein:ubiquinone oxidoreductase. These results aid in an in-depth understanding of the electron transfer process and energy metabolism involved in the nicotine oxidation and provide novel insights into nicotine catabolism in bacteria.IMPORTANCE Nicotine has been studied as a model for toxic N-heterocyclic aromatic compounds. Microorganisms can catabolize nicotine via various pathways and conserve energy from its oxidation. Although several oxidoreductases have been characterized to participate in nicotine degradation, the electron transfer involved in these processes is poorly understood. In this study, we found that 6-hydroxypseudooxynicotine dehydrogenase, a key enzyme in the hybrid pyridine and pyrrolidine pathway for nicotine degradation in Agrobacterium tumefaciens S33, utilizes EtfAB as a physiological electron acceptor. Catalyzed by the membrane-associated electron transfer flavoprotein:ubiquinone oxidoreductase, the electrons are transferred from the reduced EtfAB to coenzyme Q, which then could enter into the classic ETC. Thus, the route for electron transport from the substrate to O2 could be constructed, by which ATP can be further sythesized via chemiosmosis to support the baterial growth. These findings provide new knowledge regarding the catabolism of N-heterocyclic aromatic compounds in microorganisms.
研究领域:
微生物学
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生物化学
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分子生物学
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环境科学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF APPLIED MICROBIOLOGY
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2022
·Vol. 133
·期 5
·3161-3175
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IF 4.059
Pan Deng,Wang Xing‐Yun,Zhou Jin‐Wei,Yang Liang,Khan Abbas,Wei Dong‐Qing,Li Jun‐Jian,Jia Ai‐Qun
AimsIn the current study the anti‐virulence and anti‐biofilm activities of the cinnamic acid derivative, 3‐methoxycinnamic acid, was investigated against Agrobacterium tumefaciens.Methods and ResultsBased on the disc diffusion test and β‐galactosidase activity assay, 3‐methoxycinnamic acid was shown to interfere with the quorum sensing (QS) system of A. tumefaciens. Crystal violet staining assay, phenol‐sulfuric acid method, Bradford protein assay and confocal laser scanning microscopy (CLSM) revealed that the biofilm formation of A. tumefaciens was inhibited after the treatment of 3‐methoxycinnamic acid. Employing high‐performance liquid chromatography (HPLC) analysis of culture supernatant revealed that the production of 3‐oxo‐octanoylhomoserine lactone (3‐oxo‐C8‐HSL) decreased concentration‐dependently after treatment with 3‐methoxycinnamic acid. Swimming and chemotaxis assays also indicated that 3‐methoxycinnamic acid had a good effect on reducing the motility and chemotaxis of A. tumefaciens. In addition, the RT‐qPCR, molecular docking and simulations further demonstrated that 3‐methoxycinnamic acid could competitively inhibit the binding of 3‐oxo‐C8‐HSL to TraR and down‐regulate virulence‐related genes.Conclusions3‐Methoxycinnamic acid is proved to have good anti‐virulence and anti‐biofilm activities against A. tumefaciens.Significance and Impact of the StudyThis is the first study that investigates the anti‐virulence and anti‐biofilm activities of 3‐methoxycinnamic acid against A. tumefaciens. With its potential QS‐related virulence and biofilm inhibitory activities, 3‐methoxycinnamic acid is expected to be developed as a potent pesticide or adjuvant for the prevention and treatment of crown gall caused by A. tumefaciens.
研究领域:
微生物学
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植物病理学
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药理学
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产品关联
DOI
PubMed
高影响因子
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2022
·Vol. 194
·期 8
·3453-3467
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IF 3.094
Ye Hangyu,Li Xiaojun,Li Luyuan,Zhang Yinjun,Zheng Jianyong
Aspergillus niger has been used for homologous and heterologous expressions of many protein products. In this study, the α-L-rhamnosidase from A. niger (Rha-N1, GenBank XP_001389086.1) was homologously expressed in A. niger 3.350 by Agrobacterium tumefaciens -mediated transformation. The enzyme activity of Rha-N1 was 0.658 U/mL, which was obtained by cultivation of engineered A. niger in a 5-L bioreactor. Rha-N1 was purified by affinity chromatography and characterized. The optimum temperature and optimum pH for Rha-N1 were 60 °C and 4.5, respectively. Enzyme activity was promoted by Al 3+ , Li + , Mg 2+ , and Ba 2+ and was inhibited by Mn 2+ , Fe 3+ , Ca 2+ , Cu 2+ , and organic solvents. The result indicated that rutin was the most suitable substrate for Rha-N1 by comparison with the other two flavonoid substrates hesperidin and naringin. The transformed products of isoquercitrin, hesperetin-7-O-glucoside, and prunin were identified by LC–MS and 1 H-NMR.
研究领域:
生物化学
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分子生物学
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生物技术
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
·
2024
·Vol. 25
·期 17
·9456
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IF 4.9
Xinru Hou,Kewen Zhang,Yingmin Lyu
Oriental hybrid lilies, known for their vibrant colors, diverse flower shapes, and long blooming seasons, require annual bulb propagation in horticultural production. This necessity can lead to higher production costs and limit their use in landscaping. The LA hybrid lily ‘Aladdin’ has shown strong self-reproduction capabilities in optimal cultivation environments, producing numerous high-quality underground stem bulblets. This makes it a valuable model for studying bulblet formation in lilies under natural conditions. Through transcriptome data analysis of different developmental stages of ‘Aladdin’ bulblets, theLaLBD37gene, linked to bulblet formation, was identified. Bioinformatics analysis, subcellular localization studies, and transcriptional activation activity tests were conducted to understand the characteristics ofLaLBD37. By introducing theLaLBD37gene into ‘Sorbonne’ aseptic seedlings via Agrobacterium-mediated transformation, resistant plants were obtained. Positive plants were identified through various methods such as GUS activity detection, PCR, and fluorescence quantitative PCR. Phenotypic changes in positive plants were observed, and various physiological indicators were measured to confirm the role ofLaLBD37in bulblet formation, including soluble sugar content, starch content, sucrose synthase activity, and endogenous hormone levels. The findings suggest that theLaLBD37gene plays a significant role in promoting the development of lily bulblets, offering insights for enhancing the reproductive capacity of Oriental hybrid lilies and exploring the molecular mechanisms involved in lily bulb regeneration.
研究领域:
植物生物学
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分子遗传学
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园艺学
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DOI
高影响因子
危险品化学品经营许可证（带存储）