谷氨酸棒杆菌代谢工程高效生产L-缬氨酸北大核心CSCD

L-缬氨酸作为一种支链氨基酸,广泛应用于医药和饲料等领域。本研究借助多种代谢工程策略相结合的方法,构建了生产L-缬氨酸的微生物细胞工厂,实现了L-缬氨酸的高效生产。首先,通过增强糖酵解途径、减弱副产物代谢途径相结合的方式,强化了L-缬氨酸合成前体丙酮酸的供给;其次,针对L-缬氨酸合成路径关键酶—乙酰羟酸合酶进行定点突变,提高了菌株的抗反馈抑制能力,并利用启动子工程策略,优化了路径关键酶的基因表达水平;最后,利用辅因子工程策略,改变了乙酰羟酸还原异构酶和支链氨基酸转氨酶的辅因子偏好性,由偏好NADPH转变为偏好NADH,从而提高了L-缬氨酸的合成能力。在5L发酵罐中,最优谷氨酸棒杆菌工程菌株Corynebacterium glutamicum K020的L-缬氨酸产量、得率和生产强度分别达到了110g/L、0.51g/g和2.29 g/(L·h)。     

Nonparametric and semiparametric estimation with sequentially truncated survival data

In observational cohort studies with complex sampling schemes, truncation arises when the time to event of interest is observed only when it falls below or exceeds another random time, that is, the truncation time. In more complex settings, observation may require a particular ordering of event times; we refer to this as sequential truncation. Estimators of the event time distribution have been developed for simple left-truncated or right-truncated data. However, these estimators may be inconsistent under sequential truncation. We propose nonparametric and semiparametric maximum likelihood estimators for the distribution of the event time of interest in the presence of sequential truncation, under two truncation models. We show the equivalence of an inverse probability weighted estimator and a product limit estimator under one of these models. We study the large sample properties of the proposed estimators and derive their asymptotic variance estimators. We evaluate the proposed methods through simulation studies and apply the methods to an Alzheimer's disease study. We have developed an R package, seqTrun , for implementation of our method.     

Subcellular Distribution and Chemical Forms of Cadmium in the Medicine Food Homology Plant <i>Platycodon grandiflorum</i> (Jacq.) A.DC.

Although Platycodon grandiflorum (Jacq.) A.DC. is a renowned medicine food homology plant, reports of excessive cadmium (Cd) levels are common, which affects its safety for clinical use and food consumption. To enable its Cd levels to be regulated or reduced, it is necessary to first elucidate the mechanism of Cd uptake and accumulation in the plant, in addition to its detoxification mechanisms. This present study used inductively couple plasma-mass-spectrometry to analyze the subcellular distribution and chemical forms of Cd in different tissues of P. grandiflorum. The experimental results showed that Cd was mainly accumulated in the roots [predominantly in the cell wall (50.96%–61.42%)], and it was found primarily in hypomobile and hypotoxic forms. The proportion of Cd in the soluble fraction increased after Cd exposure, and the proportion of insoluble phosphate Cd and oxalate Cd increased in roots and leaves, with a higher increase in oxalate Cd. Therefore, it is likely that root retention mechanisms, cell wall deposition, vacuole sequestration, and the formation of low mobility and low toxicity forms are tolerance strategies for Cd detoxification used by P. grandiflorum. The results of this study provide a theoretical grounding for the study of Cd accumulation and detoxification mechanisms in P. grandiflorum, and they can be used as a reference for developing Cd limits and standards for other medicine food homology plants.     

A Meaningful Attempt: Applying Dielectric Barrier Discharge Plasma to Induce Polarization of Macrophages

Macrophage polarization plays an important role in many macrophage-related diseases. This study was designed to preliminarily explore the effects of dielectric barrier discharge (DBD) plasma on the polarization direction and cell activity of macrophages with different phenotypes (ie, M0, M1, and M2). The M1 macrophage marker inducible nitric oxide synthase (iNOS) and M2 macrophage marker cluster of differentiation 206 (CD206) were detected by western blot (WB). The effects of DBD plasma on macrophage viability were analyzed by using a cell counting kit-8 detection kit. M0, M1, and M2 macrophages exhibited a decrease in iNOS expression and an increase in CD206 expression after the DBD plasma intervention. Additionally, the decrease in macrophage viability remained non-significant after initiating the intervention. DBD plasma can promote the transformation of M0 and M1 macrophages to M2 macrophages, and can further enhance the expression of the M2 macrophage phenotype marker CD206. Our study not only demonstrates the potential therapeutic value of DBD plasma for macrophage-related diseases, but it also provides a new direction for research to improve the treatment of macrophage-related diseases. © 2023 Bioelectromagnetics Society.     

信号分子在生物脱氮中的作用及检测方法

生物脱氮是由微生物主导的地球氮循环中的重要环节之一,主要包括硝化、反硝化和厌氧氨氧化(anaerobic ammonium oxidation,anammox)等过程。在微生物联合作用下,污水中的有机氮及氨氮经一系列作用转化为氮气,这种经济高效、环境友好的处理工艺在世界范围内得到广泛应用。群体感应(quorum sensing,QS)以信号分子为媒介通过改变菌群密度和周围环境变化来调节微生物的各种行为。大量的研究已证实调控QS信号分子在生物脱氮中具有应用潜力。本文介绍了各种信号分子类型,从基因组学、实际应用等方面综述了各类信号分子以及检测方法,同时针对酰基高丝氨酸内酯(acyl homoserine lactones,AHLs)类信号分子在生物脱氮中的作用进行详细介绍。然而不足之处在于信号分子研究只是停留在实验室阶段,仅仅研究了单一信号分子对生物脱氮的影响。未来可将信号分子应用于实际污水,研究多种信号分子共同作用以及多种微生物之间的QS现象。     

库木塔格沙漠土壤细菌多样性及具有群体感应抑制活性放线菌筛选

【背景】群体感应抑制剂(quorum sensing inhibitor,QSI)作为抗生素潜在替代品,可有效降低致病菌传染性和毒性。沙漠土壤蕴藏着丰富的放线菌资源,是挖掘群体感应抑制剂的重要来源。【目的】解析库木塔格沙漠土壤细菌群落多样性,筛选并挖掘群体感应抑制活性放线菌资源。【方法】采用Illumina Nova Seq高通量测序技术揭示库木塔格沙漠土壤细菌群落组成,利用可培养方法进行土壤放线菌分离和鉴定;选用紫色杆菌CV026模型筛选群体感应抑制活性放线菌,并对其功能特性进行初步评价。【结果】Illumina Nova Seq高通量测序结果显示,样品土壤细菌涉及23门96目150属,优势菌门为变形菌门(Proteobacteria,61%)、放线菌门(Actinobacteria,28%),其中分枝杆菌属(Mycobacterium)为放线菌门最优势菌属(87.3%),其次为红球菌属(Rhodococcus,6.8%)和丙酸杆菌属(Cutibacterium,0.9%)。可培养方法共分离到108株放线菌,归属9科10属,其中优势菌属为链霉菌属(Streptomyces),占65....     

圈养大熊猫肠道乳酸菌分离及益生特性

【背景】大熊猫数量稀少、繁殖困难,在其生长过程中极易感染消化系统疾病甚至死亡。【目的】分析圈养大熊猫肠道可培养乳酸菌的群落结构与功能,筛选具有益生特性的乳酸菌菌株,为大熊猫消化系统疾病的预防和肠道微生物研究提供理论参考及菌种资源。【方法】采用3种培养基分离大熊猫粪便中的乳酸菌,通过革兰氏染色镜检和过氧化氢试验对分离的菌株进行初步鉴定,基于BOXA1R-PCR图谱遗传多样性选取代表菌株进行16S rRNA基因测序分析并进行主成分分析(principal component analysis, PCA),同时分析乳酸菌菌株安全性和益生特性。【结果】通过初步鉴定共分离获得58株乳酸菌,根据BOXA1R-PCR结果挑选20株菌进行测序,结果显示20株菌分属于明串珠菌属(Leuconostoc)、肠球菌属(Enterococcus)、魏斯氏菌属(Weissella)和链球菌属(Streptococcus)这4个属,主成分分析结果表明不同年龄段的大熊猫肠道乳酸菌群落结构存在差异。20株乳酸菌均不溶血,17株乳酸菌对11种抗生素均敏感;11株菌耐pH值2.0酸性条件,14株菌对0.3%的胆盐具有良好...     

马克斯克鲁维酵母GX-UN120糖转运蛋白基因的克隆及表征

【背景】马克斯克鲁维酵母(Kluyveromyces marxianus)具有完整的木糖代谢途径,可以高效利用木质纤维素中的木糖,因此对其糖转运蛋白基因的研究或可有效解决酵母木糖转运的相关问题。【目的】根据马克斯克鲁维酵母DMKU3-1042中KLMA＿70145和KLMA＿80101基因位点的功能预测,获得马克斯克鲁维酵母GX-UN120相应的糖转运蛋白基因序列并探究其功能。【方法】将转运蛋白基因分别克隆表达至酿酒酵母EBY.VW4000中考察重组菌株生长特性,以此间接评价对应转运蛋白的转运能力。【结果】Km＿SUT2基因编码的糖转运蛋白可有效提高宿主细胞转运木糖、阿拉伯糖、山梨糖、核糖、乳糖和葡萄糖的能力,但却不能转运甘露糖、果糖、蔗糖和半乳糖。类似地,Km＿SUT3基因编码的糖转运蛋白可提高细胞转运木糖、阿拉伯糖、山梨糖、半乳糖、核糖、乳糖和葡萄糖的能力,但却不能转运甘露糖和果糖。然而在葡萄糖存在的条件下,重组菌株对各种碳源的利用均受抑制,但Km＿SUT3转运木糖和核糖过程中受葡萄糖的抑制作用较小。【结论】马克斯克鲁维酵母GX-UN120中转运蛋白Km＿SUT2和Km＿SUT3可...     

汉阳陵陶俑表面微生物群落结构与种群多样性分析

【背景】汉阳陵是我国截至目前发现的规模最大、陪葬级别最高的汉代陶俑群,出土了大量形神毕肖的裸体俑群,考古学家认为这种裸体陶俑原本“着衣、装木臂”,称为“着衣式陶俑”,为皇室独有,异常珍贵。然而这些推测尚无科学依据,而且陶俑表面附着微生物可能会腐蚀俑体。【目的】通过比较研究汉陶俑表面微生物的组成差异,为汉裸体陶俑着有衣物且装有断臂的推测提供一定的科学佐证,同时为汉陶俑微生物腐蚀的防控提供靶标。【方法】应用微生物高通量测序与纯培养相结合的方法,对着衣式陶俑不同部位微生物进行测定。【结果】高通量测序结果显示女俑头部微生物多样性较高;男俑上半身微生物多样性最高,腿部居中,头部较低。汉陶俑表面微生物组成按俑的类别及不同部位聚集在一起,其优势菌为放线菌门的克洛斯氏菌属、糖多孢菌属、假诺卡氏菌属及链霉菌科,其他各部位优势类群相对丰度存在差异,断臂处与碳循环相关功能微生物的相对丰度最高。纯培养结果显示女俑头部的可培养细菌数量高于男俑头部可培养的细菌数量;男俑上身与腿部可培养的微生物数量高于男俑头部,表明上半身与腿部可能有衣物覆盖,导致其营养差异。【结论】汉陶俑头部与身体表面部位微生物多样性与组成存在明...