Biochemical Engineering Journal最新文献_第2页

The impact of 3D tumor spheroid maturity on cell migration and invasion dynamics 三维肿瘤球体成熟度对细胞迁移和侵袭动力学的影响

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-08 DOI: 10.1016/j.bej.2024.109567

Lingke Feng , Rong Pan , Ke Ning , Wen Sun , Yirong Chen , Yuanyuan Xie , Mingzhu Wang , Yan Li , Ling Yu

Cell motility is crucial in cancer metastasis, and understanding its regulation in tumor cells is vital for developing anti-metastatic therapies. Traditional 2D cell culture assays provide insights into cell migration but fail to replicate the complex 3D architecture of tissues in vivo. 3D cell culture models like tumor spheroids have been applied for cell migration tests. This study investigates the role of spheroid maturity in tumor cell motility, hypothesizing that spheroid maturity mirrors physiological conditions in solid tumors. Human prostate (DU 145), breast (MCF-7), and murine breast (EMT-6) cancer cells were cultured into spheroids of varying time (3, 7, and 11 days). The migration and invasion of these spheroids were analyzed, revealing that 11-day-old DU 145 spheroids demonstrated the greatest horizontal migration, correlating with RNA-seq data showing increased cell adhesion, cytoskeleton dynamics, and motility pathways. Confocal microscopy and single-cell multimode analyzer indicated higher reactive oxygen species (ROS) levels in mature spheroids, potentially activating motility pathways. Additionally, DU 145 spheroids were treated with chemotherapy reagent Doxorubicin (DOX), and the results showed that spheroids culture for 7 and 11 days exhibited greater resistance to DOX compared to spheroids cultured for 3 days. These findings highlighted the importance of considering spheroid maturity in cancer research and drug development, emphasizing the need for systematic analysis of spheroid growth conditions to ensure reproducible and reliable experimental settings.

细胞运动是癌症转移的关键，了解肿瘤细胞中的细胞运动调控对于开发抗转移疗法至关重要。传统的二维细胞培养试验可深入了解细胞迁移，但无法复制体内组织的复杂三维结构。肿瘤球体等三维细胞培养模型已被用于细胞迁移测试。本研究探讨了球体成熟度在肿瘤细胞迁移中的作用，假设球体成熟度反映了实体瘤的生理状况。研究人员将人类前列腺癌（DU 145）、乳腺癌（MCF-7）和小鼠乳腺癌（EMT-6）细胞培养成不同时间（3、7 和 11 天）的球形。对这些球形细胞的迁移和侵袭进行了分析，结果表明，培养 11 天的 DU 145 球形细胞的水平迁移能力最强，这与 RNA-seq 数据显示的细胞粘附性、细胞骨架动力学和运动途径增加有关。共聚焦显微镜和单细胞多模分析仪显示，成熟球体内的活性氧（ROS）水平较高，可能会激活运动途径。此外，用化疗试剂多柔比星（Doxorubicin，DOX）处理 DU 145 球形体，结果显示与培养 3 天的球形体相比，培养 7 天和 11 天的球形体对 DOX 的耐受性更强。这些发现突出了在癌症研究和药物开发中考虑球体成熟度的重要性，强调了对球体生长条件进行系统分析的必要性，以确保实验设置的可重复性和可靠性。

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引用次数: 0

Advanced anaerobic digestion by co-immobilization of anaerobic microbes and conductive particles in hydrogel for enhanced methane production performance 通过将厌氧微生物和导电颗粒共同固定在水凝胶中实现高级厌氧消化，从而提高甲烷生产性能

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-08 DOI: 10.1016/j.bej.2024.109563

Stella Chan , Kento Nishi , Mitsuhiko Koyama , Tatsushi Matsuyama , Junichi Ida

Recent research has increasingly focused on the enhancement of anaerobic digestion (AD) through direct interspecies electron transfer (DIET) facilitated by conductive particles (CP). Although this approach can significantly accelerate the AD process, the contact efficiency between CPs and AD microbes is relatively low due to the flow of water in a dispersed condition, leading to possible DIET inefficiency. In this study, a unique approach involving the “co-immobilization” of anaerobic microbes and multi-walled carbon nanotubes (MWCNTs) as CP into a hydrogel matrix was developed to improve the AD process. The advantages of this method include improved contact efficiency between microbes and CPs for enhanced DIET, and increased CP retention within the reactor, thereby omitting the need to compensate for CP washout. The methane production rate for the co-immobilized hydrogel was 2.5-fold and 1.9-fold faster than that of the control (dispersed sludge) and conventional DIET (dispersed sludge with MWCNT addition), respectively. Microbial analysis indicated the enrichment of functional microbes such as Anaerolineacea, Sedimentibacteraceae, Rhodocyclaceae, and Methanothrichaceae, which could be involved in the DIET under co-immobilized conditions. These results demonstrate the potential of the proposed method for realizing an advanced continuous AD process through improved DIET.

最近的研究越来越关注通过导电颗粒（CP）促进种间直接电子传递（DIET）来提高厌氧消化（AD）的效果。虽然这种方法可以大大加快厌氧消化过程，但由于水流处于分散状态，CP 与厌氧消化微生物之间的接触效率相对较低，可能导致 DIET 效率低下。本研究开发了一种独特的方法，将厌氧微生物和作为 CP 的多壁碳纳米管（MWCNTs）"共同固定 "到水凝胶基质中，以改进厌氧消化工艺。这种方法的优点包括提高微生物与氯化石蜡之间的接触效率，从而增强 DIET，并增加氯化石蜡在反应器中的保留率，从而无需补偿氯化石蜡的冲刷。与对照组（分散污泥）和传统 DIET（添加了 MWCNT 的分散污泥）相比，共固定水凝胶的甲烷生产率分别提高了 2.5 倍和 1.9 倍。微生物分析表明，Anaerolineacea、Sedimentibacteraceae、Rhodocyclaceae 和 Methanothrichaceae 等功能微生物富集，它们可能参与共同固定条件下的 DIET。这些结果表明，所提出的方法具有通过改进 DIET 实现先进的连续厌氧消化（AD）工艺的潜力。

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引用次数: 0

Developing an off-site bicarbonation absorber system to promote microalgal fixation of CO2 in exhaust gas from biogas upgrading 开发异地碳酸氢盐吸收系统，促进微藻固定沼气提纯废气中的二氧化碳

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-08 DOI: 10.1016/j.bej.2024.109558

Yulun Wu , Zhao Li , Xin Wang , Zaiyin Yu , Weiguang Mao , Cai Cheng , Guanmou Che , Jun Cheng

In order to address the risk of explosion due to CH₄ from exhaust gas produced during biogas upgrading in closed carbon fixation systems employing photosynthetic microalgae, an off-site bicarbonation absorber system was developed to promote microalgal CO₂ fixation under atmospheric pressure. The abundant CO₂ in the biogas upgrading exhaust gas (≥90 vol% CO₂, ≤10 vol% CH₄) reacted with a Na₂CO₃ solution in the off-site bicarbonation absorber to produce NaHCO₃, which was used as carbon source for microalgal growth in enclosed column photobioreactors. After the reaction, CH₄ was discharged outside the bicarbonation absorber because it did not react with the Na₂CO₃ solution and was extremely difficult to dissolve in water, thereby avoiding the explosion risk due to accumulated CH₄ in the enclosed column photobioreactors. The experimental results showed that the Spirulina growth rate first increased 1.7 times, peaking at 0.6 g/L/d, and then decreased when the bicarbonation reaction time (optimal 50 min), absorber diameter (optimal 10 cm), initial Na₂CO₃ concentration (optimal 173 mM), and exhaust gas aeration rate (optimal 100 sccm) increased. The optimal molar ratio of NaHCO₃ to total inorganic carbon in the bicarbonation absorber solution reached 79 %. The sufficient HCO₃^- supply and suitable pH of the microalgal solution improved the synthesis of photosynthetic pigments in the microalgal cells and enhanced their photochemical efficiency and carbon sequestration rates.

为了解决采用光合作用微藻的封闭式碳固定系统在沼气升级过程中产生的废气中的 CH4 导致爆炸的风险，开发了一种异地碳酸氢盐吸收系统，以促进微藻在大气压力下固定 CO2。沼气提纯废气中丰富的 CO2（≥90 vol% CO2，≤10 vol% CH4）与异位重碳酸盐吸收器中的 Na2CO3 溶液反应生成 NaHCO3，NaHCO3 用作封闭柱状光生物反应器中微藻生长的碳源。反应结束后，由于 CH4 不会与 Na2CO3 溶液发生反应，且极难溶解于水，因此 CH4 被排放到了碳酸氢盐吸收器之外，从而避免了封闭式柱状光生物反应器中累积的 CH4 所带来的爆炸风险。实验结果表明，当重碳酸化反应时间（最佳值为 50 分钟）、吸收器直径（最佳值为 10 厘米）、初始 Na2CO3 浓度（最佳值为 173 mM）和废气通气速率（最佳值为 100 sccm）增加时，螺旋藻的生长速率先增加 1.7 倍，达到 0.6 g/L/d 的峰值，然后下降。双碳酸化吸收器溶液中 NaHCO3 与总无机碳的最佳摩尔比达到 79%。微藻溶液中充足的 HCO3- 供应和适宜的 pH 值改善了微藻细胞中光合色素的合成，提高了其光化学效率和固碳率。

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引用次数: 0

A novel tubular single-chamber microbial electrolysis cell for efficient methane production from industrial potato starch wastewater 从工业马铃薯淀粉废水中高效生产甲烷的新型管式单室微生物电解池

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-07 DOI: 10.1016/j.bej.2024.109561

Gaoyuang Shang , Jinpeng Yu , Kai Cui , Hong Zhang , Yuhan Guo , Menglong Zhao , Chengjun Wang , Kun Guo

The integration of microbial electrolysis cells (MEC) with anaerobic digestion (AD) shows great promise for enhancing methane production from high-COD wastewater. However, an efficient MEC-AD reactor design remains elusive. Here, a novel tubular single-chamber MEC-AD reactor was constructed to treat potato starch wastewater (COD over 20,000 mg/L). The concentric and compact design of the stainless-steel cathode and anode reduced internal resistance, resulting in enhanced methane production. Applying −0.2 V vs. Ag/AgCl to the anode increased methane production by 1.73 times compared to the open circuit and halved hydraulic retention time. Moreover, the reactor achieved an average methane content of 82.57 %, which was 23.89 % higher than the open circuit. The reactor showed a total COD removal of 92.2 %, which was 24 % higher than the open circuit. Additionally, base consumption to maintain pH was reduced to one-sixth of that in conventional AD, preventing volatile fatty acid accumulation. Microbial analysis showed Geobacter (63.4 %) and Methanobacterium (96.8 %) were highly enriched in the anode and cathode biofilms, respectively. The proportion of fermentative bacteria also increased in the MEC-AD system. These results demonstrate the effectiveness of the tubular single-chamber MEC-AD reactor in enhancing methane production from potato starch wastewater, with strong potential for scale-up applications.

微生物电解池（MEC）与厌氧消化（AD）的结合为提高高 COD 废水的甲烷产量带来了巨大希望。然而，高效的 MEC-AD 反应器设计仍然遥遥无期。在此，我们建造了一个新型管式单室 MEC-AD 反应器来处理马铃薯淀粉废水（COD 超过 20,000 mg/L）。不锈钢阴极和阳极的同心紧凑设计降低了内阻，从而提高了甲烷产量。在阳极上施加 -0.2 V 对 Ag/AgCl 的电压，甲烷产量比开式回路增加了 1.73 倍，水力停留时间缩短了一半。此外，反应器的平均甲烷含量达到 82.57%，比开式回路高出 23.89%。反应器的化学需氧量总去除率为 92.2%，比开式回路高 24%。此外，维持 pH 值所需的碱消耗量减少到传统厌氧消化法的六分之一，从而防止了挥发性脂肪酸的积累。微生物分析表明，阳极生物膜和阴极生物膜中分别高度富集了 Geobacter（63.4%）和 Methanobacterium（96.8%）。在 MEC-AD 系统中，发酵菌的比例也有所增加。这些结果证明了管式单室 MEC-AD 反应器在提高马铃薯淀粉废水甲烷产量方面的有效性，具有很强的放大应用潜力。

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引用次数: 0

Improvement of sludge dewatering performance by persulfate advanced oxidation combined with LDH: Synergistic effect of free radical and non-free radical and reuse of deep-dewatered sludge cake 过硫酸盐高级氧化结合 LDH 改善污泥脱水性能：自由基和非自由基的协同效应以及深度脱水污泥饼的再利用

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-07 DOI: 10.1016/j.bej.2024.109560

Jingyu Lv, Lei Song, Yang Guo

The high-water content of sludge in wastewater plant will influence the transportation and utilization. In this study, a new method for improving sludge dewatering by pyrite (FeS₂) activated persulfate (PMS) combined with layered double hydroxide (LDH) was proposed. After conditioning, the water content (Wc) and specific resistance (SRF) of sludge decreased from 97.12 % and 1.83 × 10¹³ m/kg to 71.39 % and 1.84 × 10¹² m/kg, severally. SEM and particle size analysis showed the system could destroy sludge cells effectively.The mechanism analysis of protein and polysaccharide content, 3D-EEM, FTIR, XPS results showed that FeS₂/PMS-LDH combined system was beneficial to break down the sludge extracellular polymer (EPS), transform and accumulate the organic matter into the EPS outer layer, release the bound water. Both free radical and non-free radical play a role in oxidation, and they cooperate to break EPS. The effective phosphate adsorption performance of the biochar adsorbent prepared from dehydrated sludge cake was also investigated. The adsorption behavior of phosphate on biochar from dewatered sludge cake belongs to uniform chemical monolayer adsorption. When T = 298k, PH = 5, the maximum adsorption capacity is 20.255 mg/g. The introduction of LDH is helpful to enhance the sludge dewatering and the adsorption of phosphate. To sum up, the combined conditioning method considers the effectiveness of sludge dewatering and the feasibility of sludge cake disposal and utilization.

污水厂污泥含水率高，会影响污泥的运输和利用。本研究提出了一种利用黄铁矿（FeS2）活化过硫酸盐（PMS）结合层状双氢氧化物（LDH）改善污泥脱水的新方法。经过调理后，污泥的含水率（Wc）和比阻力（SRF）分别从 97.12 % 和 1.83 × 1013 m/kg 降至 71.39 % 和 1.84 × 1012 m/kg。蛋白质和多糖含量的机理分析、3D-EEM、傅立叶变换红外光谱、XPS 结果表明，FeS2/PMS-LDH 组合系统有利于分解污泥胞外聚合物（EPS），将有机物转化并积聚到 EPS 外层，释放结合水。自由基和非自由基都在氧化过程中发挥作用，它们共同作用使 EPS 断裂。研究还考察了脱水污泥饼制备的生物炭吸附剂对磷酸盐的有效吸附性能。脱水污泥饼生物炭对磷酸盐的吸附行为属于均匀化学单层吸附。当 T = 298k，PH = 5 时，最大吸附容量为 20.255 mg/g。LDH的引入有助于提高污泥脱水和磷酸盐的吸附能力。总之，联合调理法既考虑了污泥脱水的有效性，又考虑了污泥饼处置和利用的可行性。

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引用次数: 0

Sustainable remediation of piggery wastewater using a novel mixotrophic Chlorella sorokiniana Cbeo for high value biomass production 利用新型混养小球藻 Cbeo 对养猪场废水进行可持续修复，以生产高价值生物质

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-06 DOI: 10.1016/j.bej.2024.109555

Do Thi Cam Van , Dang Thi Mai , Bui Thi Thu Uyen , Nguyen Thi Phuong Dung , Lu Thi Thu Ha , Nguyen Thi Lieu , Dang Nhat Minh , Tran Dang Thuan , Le Truong Giang

Piggery wastewater (PW) contains high density of organic carbon (COD), nitrogen (NH₄⁺-N and TN) and phosphorous (TP), which are essential nutrients for microalgae growth. This work was attempted to use a newly isolate Chlorella sorokiniana C_beo for recovery these compounds into its biomass via mixotrophic cultivation. Critical factors including level of ammonia, C/N ratio, pH, light intensity, sterilized/unsterilized media, and indoor/outdoor cultivations affecting biomass production and nutrients removal efficiencies were investigated. Data revealed that C. sorokiniana C_beo achieved the optimal growth in the unsterilized medium at NH₄⁺-N concentration, C/N ratio, initial pH, and light intensity of 250 mg/L, 10/1, 7, and 150 μmol/m²/s, respectively. Under the optimal conditions, dry cell weight (DCW) reached the maximal level of 4.30 g/L, which was slightly higher than 4.14 g/L determined for the sterilized medium. In 30 L-scale photobioreactor, C. sorokiniana C_beo grown under indoor and outdoor achieved DCW of 3.61 and 3.19 g/L, respectively. COD, NH₄⁺-N, TN, TP removal efficiencies for both conditions were determined as 91.9–96.7, 96.6–99.7, 96.2–96.4, and 98.2–100 %, respectively. The C. sorokiniana C_beo biomass contained 14–27 % lipid, 25–32 % carbohydrate, 44–48 % protein, and 0.25–0.97 % lutein. Interestingly, α-Linolenic acid (C18:3n3) was 19.84 –27.0 % of the total fatty acids. C. sorokiniana C_beo is the promising algal strain for development of a sustainable biorefinery of PW.

养猪场废水（PW）中含有高浓度的有机碳（COD）、氮（NH4+-N 和 TN）和磷（TP），这些都是微藻生长所必需的营养物质。本研究试图利用一种新分离的小球藻 Cbeo，通过混养培养将这些化合物回收到其生物量中。研究了影响生物量生产和营养物质去除效率的关键因素，包括氨水平、C/N 比、pH 值、光照强度、灭菌/未灭菌培养基以及室内/室外培养。数据显示，当 NH4+-N 浓度、C/N 比、初始 pH 值和光照强度分别为 250 mg/L、10/1、7 和 150 μmol/m2/s 时，C. sorokiniana Cbeo 在未灭菌培养基中达到最佳生长。在最佳条件下，干细胞重量（DCW）达到最大值 4.30 g/L，略高于灭菌培养基的 4.14 g/L。在 30 L 规模的光生物反应器中，在室内和室外条件下生长的 C. sorokiniana Cbeo 的干细胞重量分别为 3.61 和 3.19 g/L。两种条件下的 COD、NH4+-N、TN、TP 去除率分别为 91.9%-96.7%、96.6%-99.7%、96.2%-96.4% 和 98.2%-100%。C. sorokiniana Cbeo 的生物质含有 14-27 % 的脂质、25-32 % 的碳水化合物、44-48 % 的蛋白质和 0.25-0.97 % 的叶黄素。有趣的是，α-亚麻酸（C18:3n3）占脂肪酸总量的 19.84 -27.0%。C. sorokiniana Cbeo 是开发 PW 可持续生物精炼厂的理想藻种。

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引用次数: 0

Experimental determination of maximum shear stress in Mobius® Breez perfusion microbioreactors and comparative analysis with stirred tank bioreactors Mobius® Breez 灌注微型生物反应器中最大剪切应力的实验测定以及与搅拌罐生物反应器的比较分析

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-11-06 DOI: 10.1016/j.bej.2024.109556

Patrick Romann , Dan Trunov , Ondřej Šrom , Harry L.T. Lee , Kevin S. Lee , Ryan Trocki , David Ephraim , Jean-Marc Bielser , Jonathan Souquet , Miroslav Šoóš , Thomas K. Villiger

Perfusion processes have experienced increased popularity in recent years due to their ability to sustain high cell densities and productivities in biopharmaceutical production, offering advantages over traditional batch and fed-batch cultivation methods. The Mobius® Breez microbioreactor significantly reduces experimental effort by downsizing the classical volume of perfusion bioreactors to the mL range and thus represents a valuable tool for process development. However, miniaturization has raised questions regarding comparability with traditional bioreactors in terms of the physical environment, such as hydrodynamic shear stress. Therefore, the maximum hydrodynamic shear stress, cultivation performance, and membrane-wall contact were evaluated to elucidate the system's behavior. Findings reveal two distinct operational conditions, distinguished by the presence or absence of membrane-wall contact, resulting in varying levels of hydrodynamic stress. Conditions lacking membrane contact demonstrate stress levels within safe operating thresholds for CHO cells, while those involving membrane contact exceed these thresholds, potentially leading to cell damage. Through the identification of critical frequencies of membrane motion, this study offers insights for optimizing microbioreactor operation and enhancing overall bioprocess efficiency.

灌流工艺近年来越来越受欢迎，因为它能够在生物制药生产中维持较高的细胞密度和生产率，与传统的间歇式和喂料式培养方法相比更具优势。Mobius® Breez 微型生物反应器将传统灌流生物反应器的体积缩小到毫升范围，大大减少了实验工作量，因此是工艺开发的重要工具。然而，微型化在物理环境（如流体动力剪切应力）方面引起了与传统生物反应器可比性的问题。因此，我们对最大流体动力剪切应力、培养性能和膜壁接触进行了评估，以阐明该系统的行为。研究结果显示了两种不同的操作条件，以膜壁接触的有无来区分，从而产生不同程度的流体动力应力。没有膜接触的情况下，应力水平在 CHO 细胞的安全操作阈值范围内，而有膜接触的情况下，应力水平超过了这些阈值，有可能导致细胞损伤。通过确定膜运动的临界频率，这项研究为优化微生物反应器的运行和提高整体生物处理效率提供了启示。

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引用次数: 0

Development of inducible packaging cell line for rAAV production via CRISPR-Cas9 mediated site-specific integration 通过 CRISPR-Cas9 介导的位点特异性整合，开发用于生产 rAAV 的可诱导包装细胞系

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-10-31 DOI: 10.1016/j.bej.2024.109552

Qiang Fu , Yongdan Wang , Emily Doleh , Mark Blenner , Seongkyu Yoon

AAV-mediated gene therapy is a quickly growing segment of the pharmaceutical market; however, the current transient transfection process to produce rAAV has several challenges. The stable cells are ideal for large-scale continuous production, overcoming the drawbacks in the current transient transfection and streamlining rAAV production. In this study, we proposed to use synthetic inducible promoters to control the viral component expression and develop the baseline of HEK293T stable cells via site-specific integration mediated with CRISPR-Cas9, targeting safe harbor sites of human genome (ROSA26, AAVS1, and CCR5 locus). With a total of three round integrations, stable cell pools were developed and evaluated at each round of integration. Single clones were further characterized for each integration round. Regarding the stable pools, the 5’ and 3’ junction PCR results confirmed the site-specific integration to each locus. The genome copy result showed that AAV components, including Rep78/68, E2A, E4orf6, Cap, and Rep52/40, were successfully integrated into the host cell genome. Genome and capsid titer after induction confirmed rAAV production for stable cell pools in each round. The packaging cell line (after 2nd round integration) was able to produce rAAV. However, it was observed that the genome titer was ten-fold lower than that of rAAV products done with triple plasmids transfection. The out-to-out PCR and qPCR assay results further confirm the site-specific integration. This research demonstrates the feasibility of developing the inducible stable cell line with the refactored viral vectors via a site-specific integration.

AAV 介导的基因疗法是医药市场中一个快速增长的细分市场；然而，目前生产 rAAV 的瞬时转染工艺面临着一些挑战。稳定细胞是大规模连续生产的理想选择，它克服了目前瞬时转染的缺点，简化了 rAAV 的生产过程。在本研究中，我们提出使用合成诱导启动子来控制病毒成分的表达，并通过 CRISPR-Cas9 介导的位点特异性整合，以人类基因组的安全港位点（ROSA26、AAVS1 和 CCR5 位点）为目标，开发 HEK293T 稳定细胞基线。总共进行了三轮整合，在每一轮整合中都开发并评估了稳定细胞池。对每轮整合的单克隆进行了进一步鉴定。关于稳定细胞池，5'和 3'连接PCR结果证实了每个基因座的特异性整合。基因组拷贝结果显示，包括 Rep78/68、E2A、E4orf6、Cap 和 Rep52/40 在内的 AAV 成分已成功整合到宿主细胞基因组中。诱导后的基因组和囊壳滴度证实，每一轮都有稳定的细胞池产生了 rAAV。包装细胞系（第二轮整合后）能够产生 rAAV。但观察到基因组滴度比三重质粒转染的 rAAV 产物低十倍。从外到内的 PCR 和 qPCR 检测结果进一步证实了位点特异性整合。这项研究证明了通过位点特异性整合利用重构病毒载体开发可诱导的稳定细胞系的可行性。

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引用次数: 0

Impact of tetracycline on mixotrophic denitrification process under different sulfur to nitrogen ratios 不同硫氮比条件下四环素对混养反硝化过程的影响

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-10-31 DOI: 10.1016/j.bej.2024.109557

Bohan Lv , Yang-Guo Zhao , Yue Chen , Mupindu Progress , Mengchun Gao , Liang Guo , Junyuan Ji , Chunji Jin

The sulfur-based autotrophic-heterotrophic denitrification, i.e., mixotrophic denitrification, is suitable for the nitrate and antibiotics removal in aquaculture tailwater at a low COD to nitrogen (C/N) ratio. This study focused on the effect of tetracycline (TC) on mixotrophic denitrification under different S/N ratios. Two bioreactors were simultaneously operated with or without dosing tetracycline under different sulfur to nitrogen (S/N) ratios of 3.94, 4.64 and 5.94. The results showed that the removal rate of total inorganic nitrogen (TIN) increased from 0.25 to 0.69 mg N L⁻¹ min⁻¹ with the rise of S/N ratio, while TC dosage significantly declined the removal efficiency of TIN. Dissimilatory nitrogen reduction to ammonia (DNRA) bacteria was detected when exposing to TC, indicating that DNRA presented more resistance to TC. The removal efficiency of TC in the denitrification system reached the maximum of 22.87 % at S/N of 4.64. Meanwhile the genus Marinicella was detected at this phase, which was conducive to the degradation of organic pollutants. This study found that TC promoted the accumulation of ammonia nitrogen, and had a great effect on sulfur autotrophic bacteria at S/N of 5.94. The removal of TC mainly depended on microbial co-metabolism, and there was a significant correlation between the reduction of TC concentration and the decrease of sulfur compounds (p < 0.05). 4.64 is the best S/N ratio for the mixotrophic denitrification process, which revealed maximum nitrate and TC removal rates.

以硫为基础的自养-异养反硝化作用，即混养反硝化作用，适用于在低化学需氧量（COD）-氮（C/N）比条件下去除养殖尾水中的硝酸盐和抗生素。本研究主要探讨了四环素（TC）在不同信噪比条件下对混养反硝化作用的影响。在不同的硫氮（S/N）比（3.94、4.64 和 5.94）条件下，同时运行两个生物反应器，是否投加四环素。结果表明，随着硫氮比的升高，总无机氮（TIN）的去除率从 0.25 mg N L-1 min-1 提高到 0.69 mg N L-1 min-1，而四环素的投加量则显著降低了总无机氮的去除效率。在暴露于 TC 的情况下，检测到了分解氮还原氨（DNRA）细菌，这表明 DNRA 对 TC 有更强的抵抗力。反硝化系统对 TC 的去除率在信噪比为 4.64 时达到最高，为 22.87%。同时，在这一阶段检测到了马林杆菌属，这有利于有机污染物的降解。本研究发现，在信噪比为 5.94 时，TC 促进了氨氮的积累，并对硫自养菌有很大影响。TC 的去除主要依赖于微生物的协同代谢，TC 浓度的降低与硫化合物的减少之间存在显著相关性（p <0.05）。4.64 是混养反硝化过程的最佳信噪比，它显示了最大的硝酸盐和 TC 去除率。

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引用次数: 0

Pros and cons of airlift and bubble column bioreactors: How internals improve performance 气浮和气泡柱生物反应器的优缺点：内部结构如何提高性能

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biochemical Engineering Journal

Pub Date : 2024-10-30 DOI: 10.1016/j.bej.2024.109539

Carolin Bokelmann , Jason Bromley , Ralf Takors

Gas fermentation is a promising technology of high commercial interest, particularly for capturing CO₂ and CO from industrial off-gases to reduce greenhouse gas emissions and replace fossil fuels for bulk chemical production. Therefore, evaluating promising bioreactor settings ab initio is a crucial step. Whereas alternate configurations may be tested in laborious scale up studies, the procedure may be accelerated by in silico studies that accompany or even partially replace wet-lab work once the models are validated. In this context, the current study compares various pneumatically agitated reactor types – bubble column reactor (BCR), annulus- and center-rising internal-loop airlift reactor (AR-IL-ALR and CR-IL-ALR), and external-loop airlift reactor (EL-ALR) – to identify advantages and disadvantages for the given application based on computational fluid dynamics (CFD) models. Process performance is optimized by introducing internal structures to guide the flow. Despite a significant increase in the mass transfer coefficient (

k_{L} a

) through internal modifications, the CR-IL-ALR still exhibited the poorest performance. The optimized AR-IL-ALR demonstrated good mixing and, after introducing an open-cone shaped internal in the head part and a conical bottom, superior mass transfer, achieving an enhancement over 10 % in the mass transfer coefficient to 315 1/h. This study thereby outlines the potential of internal structures for process improvement, as well as the value of a priori in silico design of reactor configurations.

气体发酵是一项极具商业价值的技术，特别是从工业废气中捕获二氧化碳和一氧化碳，以减少温室气体排放，并取代化石燃料用于大宗化学品生产。因此，对有前景的生物反应器设置进行初始评估是至关重要的一步。虽然可以在费力的放大研究中测试替代配置，但在模型得到验证后，可以通过伴随甚至部分取代湿实验室工作的硅学研究来加速这一过程。在这种情况下，当前的研究比较了各种气动搅拌反应器类型--气泡塔反应器（BCR）、环形和中心上升内循环气动反应器（AR-IL-ALR 和 CR-IL-ALR）以及外循环气动反应器（EL-ALR）--根据计算流体动力学（CFD）模型确定特定应用的优缺点。通过引入内部结构来引导气流，从而优化工艺性能。尽管通过内部改造大大提高了传质系数（kLa），但 CR-IL-ALR 的性能仍然最差。优化后的 AR-IL-ALR 不仅混合效果良好，而且在头部引入开放式锥形内部结构和锥形底部后，传质效果更佳，传质系数提高了 10% 以上，达到 315 1/h。这项研究由此概述了内部结构在改进工艺方面的潜力，以及对反应器配置进行先验硅设计的价值。

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引用次数: 0