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基于细菌体系生产双链RNA的条件优化
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作者 计慧君 林羿光 +1 位作者 付彤煜 郑思春 《环境昆虫学报》 CSCD 北大核心 2023年第3期703-710,共8页
化学杀虫剂是农业害虫防治的主要手段,然而农业害虫已对化学杀虫剂产生了抗药性。RNA农药的应用将是减少使用化学杀虫剂的一种途径。RNA农药具有专一、高效、易降解和对环境友好等优点而受到了关注,但在靶标分子、靶标分子的双链RNA(dsR... 化学杀虫剂是农业害虫防治的主要手段,然而农业害虫已对化学杀虫剂产生了抗药性。RNA农药的应用将是减少使用化学杀虫剂的一种途径。RNA农药具有专一、高效、易降解和对环境友好等优点而受到了关注,但在靶标分子、靶标分子的双链RNA(dsRNA)生产工艺和应用制剂等方面仍需进一步研究。本研究以褐飞虱Nilaparvata lugens蛋白激酶B基因的dsRNA(dsNlAKT)为例,对利用细菌体系生产dsRNA的方法进行了探究。将NlAKT构建进L4440载体后,将其转化到缺乏核酸酶(RNaseⅢ)的大肠杆菌Escherichia coli HT115(DE3)中,进行了dsRNA生产条件的测试。结果表明:(1)当异丙基-β-D-硫代半乳糖苷(IPTG)工作浓度为0.1 mM或0.5 mM时dsRNA的产量没有明显变化,而当其工作浓度增加至1 mM时dsRNA产量明显减少;(2)在IPTG诱导时间为2~8 h范围内,添加IPTG后诱导6 h获得的dsRNA产量最多;(3)在实验室常规提取RNA方法中,增加低剂量溶菌酶预处理这一步骤可增加提取产物中dsRNA的含量。试验结果证明了采用工作浓度为0.1 mM的IPTG诱导6 h的生产条件,并在提取过程中增加溶菌酶的预处理步骤有助于获得较高的dsRNA含量。研究结果为RNA农药的生产条件提供了实验依据。 展开更多
关键词 双链RNA RNA农药生产 细菌体系 RNA提取
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Study on the Quantity Dynamic Changes of Heterobacteria and Vibrios in Larvae Industrialized Culture System 被引量:7
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作者 马妍 李健 +2 位作者 王斌 王群 刘淇 《Agricultural Science & Technology》 CAS 2008年第2期116-121,共6页
[ Objective ] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp. [ Method ] The heterobacteria, vibr... [ Objective ] The dynamic change of heterobacteria and vibrios in larvae industrialized culture system was studied to provide scientific reference for healthy cultivation of shrimp. [ Method ] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were monitored in larvae industrialized culture system. [ Result] The heterobacteria, vibrios and pathogenic vibrio parahaemolyticus were the most in fertilized eggs of shrimp but the least in nauplius, then their number would increase with growth. During whole rearing period, both boterobacteria in larvae, vibrios in water would increase by one order of magnitude, while both vibrios in larvae and heterobacteria in water would increase by two orders of magnitude. There were many heterobacteria and vibrios but few vibrio parahaemolyticus in living bait. The correlation coefficients between larvae and heterobacteria and vibrios in water were 0. 704 and 0. 840 in culture system respectively, while the correlation among heterobacteria, vibrios in living bait and larvae, water were weak or negative. [ Conclusion ] There was a dynamic relation between water and larvae in rearing period, and restrictly control of culture condition would restrain the occurrence of disease caused by vibrio parahaemolyticus, besides that bacteria number in bait was not obviously correlated with bacteria nubmer in culture system. 展开更多
关键词 Chinese shrimp larvae Heterotrophic bacteria VIBRIO Vibrio parahaemolyticus Dynamic change
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Cultural and luminescent Conditions of a Marine luminous Bacterium
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作者 方宏达 董燕红 +2 位作者 袁茵 李秀芹 易斌 《Marine Science Bulletin》 CAS 2008年第1期46-53,共8页
Study of marine noctilucence in marine is important to fishery, environmental monitoring and military affairs. A luminous bacterial strain D2 was isolated from the marine sediment samples collected near Donghai Island... Study of marine noctilucence in marine is important to fishery, environmental monitoring and military affairs. A luminous bacterial strain D2 was isolated from the marine sediment samples collected near Donghai Island in Zhanjiang, China. The primary cultural and luminescent conditions of luminous bacterium D2 which was identified as Vibrio sp. were determined in liquid culture. The results showed that pH 7.0, 35 ℃, with 2.0 % NaCI, were the best growth conditions, and pH5 - 6, 20℃, OD600 0.08, with 3.0 % NaCI, were the optimal luminescent conditions. 展开更多
关键词 Luminous bacterium growth conditions luminescent conditions quorum sensing system
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细菌细胞表面展示体系提呈人源诺如病毒(GⅡ.4)抗原效果的评价 被引量:1
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作者 竺晖 王楠 +4 位作者 鲁飞凤 张子蕾 唐峰 徐婷 王大鹏 《病毒学报》 CAS CSCD 北大核心 2020年第3期439-445,共7页
人源诺如病毒(Human noroviruses,HuNoVs)是重要的肠胃炎病毒之一,造成了全球巨额经济负担,故其防控成为研究热点。探究不同形式的免疫原对机体体液和粘膜免疫的激发效果,成为HuNoVs疫苗研发的一个突破口。本研究利用前期以细菌细胞表... 人源诺如病毒(Human noroviruses,HuNoVs)是重要的肠胃炎病毒之一,造成了全球巨额经济负担,故其防控成为研究热点。探究不同形式的免疫原对机体体液和粘膜免疫的激发效果,成为HuNoVs疫苗研发的一个突破口。本研究利用前期以细菌细胞表面展示体系构建的假HuNoVs(GⅡ.4)和纯化的P(GⅡ.4)蛋白为对象,乳化后每隔两周分别进行Balb/C小鼠皮下多点和口服免疫,并以间接酶联免疫吸附试验测定血清中抗体的消长规律。同时,以磷酸盐缓冲液作为对照。抗体效价测定结果显示:皮下免疫组的假HuNoVs(GⅡ.4)可以有效激发小鼠体液免疫,其效价稍低于以P(GⅡ.4)蛋白作为抗原免疫获得的抗体。经过5次免疫后,血清中IgG抗体效价基本趋于稳定。但是,假HuNoVs(GⅡ.4)和P(GⅡ.4)蛋白的口服免疫组血清中均未测到IgA和IgG抗体。因此,细菌细胞表面展示体系假HuNoVs(GⅡ.4)可以作为HuNoVs疫苗候选抗原的提呈体系,为该病毒的疫苗研发提供借鉴。 展开更多
关键词 人源诺如病毒(HuNoVs) 细菌细胞表面展示体系 P蛋白 免疫 抗原提呈
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Construction of cDNA Library from Intestine, Mesentery and Coelomocyte of Apostichopus japonicus Selenka Infected with Vibrio sp. and a Preliminary Analysis of Immunity-Related Genes 被引量:1
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作者 LIU Hongzhan ZHENG Fengrong +1 位作者 SUN Xiuqin CAI Yimei 《Journal of Ocean University of China》 SCIE CAS 2012年第2期187-196,共10页
The aquaculture of sea cucumber Apostichopus japonicus (Echinodermata, Holothuroidea) has grown rapidly during recent years and has become an important sector of the marine industry in Northern China. However, with th... The aquaculture of sea cucumber Apostichopus japonicus (Echinodermata, Holothuroidea) has grown rapidly during recent years and has become an important sector of the marine industry in Northern China. However, with the rapid growth of the industry and the use of non-standard culture techniques, epidemic diseases of A. japonicus now pose increasing problems to the industry. To screen the genes with stress response to bacterial infection in sea cucumber at a genome wide level, we constructed a cDNA library from A. japonicus Selenka (Aspidochirotida: Stichopodidae) after infecting them with Vibrio sp. for 48 h. Total RNA was extracted from the intestine, mesentery and coelomocyte of infected sea cucumber using Trizol and mRNA was isolated by Oligotex mRNA Kits. The ligated cDNAs were transformed into DH5α, and a library of 3.24×105 clones (3.24×105 cfu mL-1) was obtained with the sizes of inserted fragments ranging from 0.8 to 2.5 kb. Sequencing the cDNA clones resulted in a total of 1106 ESTs that passed the quality control. BlastX and BlastN searches have identified 168 (31.5%) ESTs sharing significant homology with known sequences in NCBI protein or nucleotide databases. Among a panel of 25 putative immunity-related genes, serum lectin isoform, complement component 3, complement component 3-like genes were further studied by real-time PCR and they all increased more than 5 fold in response to Vibrio sp. challenge. Our library provides a valuable molecular tool for future study of invertebrate immunity against bacterial infection and our gene expression data indicates the importance of the immune system in the evolution and development of sea cucumber. 展开更多
关键词 Apostichopusjaponicus cDNA library expressed sequence tags immunity-related genes real-time PCR
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Characterization and in-vivo evaluation of potential probiotics of the bacterial flora within the water column of a healthy shrimp larviculture system 被引量:12
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作者 薛明 梁华芳 +1 位作者 何瑶瑶 温崇庆 《Chinese Journal of Oceanology and Limnology》 SCIE CAS CSCD 2016年第3期484-491,共8页
A thorough understanding of the normal bacterial flora associated with shrimp larviculture systems contributes to probiotic screening and disease control. The bacterial community of the water column over a commercial ... A thorough understanding of the normal bacterial flora associated with shrimp larviculture systems contributes to probiotic screening and disease control. The bacterial community of the water column over a commercial Litopenaeus vannamei larval rearing run was characterized with both culture-dependent and culture-independent methods. A total of 27 phylotypes at the species level were isolated and identified based on 16S rDNA sequence analysis. Denaturing gradient gel electrophoresis (DGGE) analysis of the V3-V5 region of 16S rRNA genes showed a dynamic bacterial community with major changes occurred from stages zoea to mysis during the rearing run. The sequences retrieved were affiliated to four phyla, Proteobacteria, Actinobacteria, Bacteroidetes, and Firmicutes, with the family Rhodobacteraceae being the most frequently recovered one. Subsequently, 13 representative strains conferred higher larval survival than the control when evaluated in the in-vivo experiments; in particular, three candidates, assigned to Phaeobacter sp., Arthrobacter sp., and Microbacteriurn sp., significantly improved larval survival (P〈0.05). Therefore, the healthy shrimp larviculture system harbored a diverse and favorable bacterial flora, which contribute to larval development and are of great importance in exploiting novel probiotics. 展开更多
关键词 Bacterial flora Litopenaeus vannamei LARVICULTURE PROBIOTICS 16S rRNA gene
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Isolation and Characterization of Heterotrophic Nitrifying Strain W1 被引量:5
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作者 吕永康 王荀 +2 位作者 刘博凯 刘玉香 杨晓华 《Chinese Journal of Chemical Engineering》 SCIE EI CAS CSCD 2012年第5期995-1002,共8页
In a high concentration substrate medium, a heterotrophic bacterium with high removal efficiency of ammonium, named W1, was isolated from activated sludge of coking wastewater treatment facility. The bacterium was Gra... In a high concentration substrate medium, a heterotrophic bacterium with high removal efficiency of ammonium, named W1, was isolated from activated sludge of coking wastewater treatment facility. The bacterium was Gram-negative, rod-shaped, and identified preliminarily as Alcaligenes sp. according to its morphological and physiological properties and its 16S rRNA gene sequence analysis. In the high concentration ammonium medium (400 mg·L 1 4 NH -N), the effects of C source, N source, C/N ratio and initial pH of medium on ammonium removal were investigated in order to determine the optimal condition for strain W1. The maximum ammonium removal was around 95% in 4 days in an improved medium. The production of N 2 gas was examined in a closed system that was full of pure oxygen at the beginning. N 2 gas was detected in the system after 4 days of cultivation, which further testified that strain W1 has heterotrophic nitrification and aerobic denitrification abilities simultaneously. 展开更多
关键词 heterotrophic nitrification aerobic de-nitrification high concentration ammonium Alcaligenes sp.
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Collective motion of bacteria and their dynamic assembly behavior
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作者 冯靖靖 何彦 《Science China Materials》 SCIE EI CSCD 2017年第11期1079-1092,共14页
In recent years, active matter systems have attracted considerable attentions due to their complex dynamic behaviors in physical and material science. In particular, microorganism systems have served as model systems ... In recent years, active matter systems have attracted considerable attentions due to their complex dynamic behaviors in physical and material science. In particular, microorganism systems have served as model systems for observing dynamic assembly and collective motility of active particles and significant progresses have been made on in-depth understanding of how high density bacteria colony behaves in the non-equilibrium state. In this mini-review, we mainly focus on the collective motion of bacteria and their dynamic assembly from four aspects: (1) the general phenomenon and biological mechanism of bacterial collective motion; (2) the common experimental techniques for studying bacterial motility; (3) some active systems on exploring bacterial collective behavior, which include both non-restricted free suspensions and those in relative confined geometric space; (4) the phenomenological and descriptive statistical methods and physical models on the underlying laws that lead to large-scale coordinate patterns in multicellular systems. This review aims to give a general picture of the collective motion in bacterial active matter systems experimentally and theoretically in order to reflect the interplays between individuals among populations in motion. It is expected that the general regulation rules related to the boundary effects in the complex systems and materials can be elucidated to some extent. 展开更多
关键词 active matter bacterial swarming collective behav-ior
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