Journal of Microbiology

Journal Articles

Transposon insertion site sequencing (TIS) of Pseudomonas aeruginosa: Hongbaek Cho; J. Microbiol. 2021;59(12):1067-1074. Published online December 4, 2021; DOI: https://doi.org/10.1007/s12275-021-1565-y

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Transposon insertion site sequencing (TIS) is a technique that determines the insertion profile of a transposon mutant library by massive parallel sequencing of transposon-genomic DNA junctions. Because the transposon insertion profile reflects the abundance of each mutant in the library, it provides information to assess the fitness contribution of each genetic locus of a bacterial genome in a specific growth condition or strain background. Although introduced only about a dozen years ago, TIS has become an important tool in bacterial genetics that provides clues to study biological functions and regulatory mechanisms. Here, I describe a protocol for generating high density transposon insertion mutant libraries and preparing Illumina sequencing samples for mapping the transposon junctions of the transposon mutant libraries using Pseudomonas aeruginosa as an example.

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Optimizing phage-based mutant recovery and minimizing heat effect in the construction of transposon libraries in Staphylococcus aureus
Sally W. Yousief, Nader Abdelmalek, Bianca Paglietti
Scientific Reports.2024;[Epub] CrossRef
The biological essence of synthetic lethality: Bringing new opportunities for cancer therapy
Meiyi Ge, Jian Luo, Yi Wu, Guobo Shen, Xi Kuang
MedComm – Oncology.2024;[Epub] CrossRef
Optimization of Transposon Mutagenesis Methods in Pseudomonas antarctica
Sangha Kim, Changhan Lee
Microorganisms.2023; 11(1): 118. CrossRef
Construction of high-density transposon mutant library of Staphylococcus aureus using bacteriophage ϕ11
Wonsik Lee
Journal of Microbiology.2022; 60(12): 1123. CrossRef

RNA Interference Targeting Nucleocapsid Protein Inhibits Porcine Reproductive and Respiratory Syndrome Virus Replication in Marc-145 Cells: Minnan Yang , Qun Xiang , Xiaodong Zhang , Xiang Li , Seydou Sylla , Zhuang Ding; J. Microbiol. 2014;52(4):333-339. Published online March 29, 2014; DOI: https://doi.org/10.1007/s12275-014-3419-3

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Abstract

Porcine reproductive and respiratory syndrome (PRRS) is an important disease, which leads to severe economic losses in swine-producing areas of the world. However, current antiviral strategies cannot provide highly effective protection. In this study, three theoretically effective interference target sites (71-91, 144-164, 218-238) targeting the nucleocapsid (N) gene of PRRSV were designed and selected, and then three siRNA-expressing plasmids were constructed, respectively named p2.1-N71, p2.1-N144, and p2.1-N218. The recombinant siRNA-expressing plasmids were transfected into Marc-145 cells; then the cells were infected with PRRSV (JL07SW strain); finally, after incubation for 48 h, the antiviral activity of those siRNA-expressing plasmids in Marc-145 cells was assessed by cytopathic effects, virus titers, indirect immunofluorescence, and quantitative real-time PCR. Experimental results demonstrated that these three siRNA-expressing plasmids could effectively and significantly inhibit the replication of PRRSV by 93.2%, 83.6%, and 89.2% in Marc-145 cells, respectively. Among these three siRNA-expressing plasmids, p2.1-N71 was found to be most effective, while p2.1-N144 and p2.1-N218 displayed relatively weak inhibition of virus replication. The results indicated that siRNA-expressing plasmids targeting the N gene of PRRSV could significantly inhibit PRRSV replication in Marc-145 cells. Based on our experimental results and previous reports, the 71-91, 179-197, and 234-252 sites of the N gene are good choices to effectively inhibit the replication of PRRSV, and this RNA interference technique can be a potential anti-PRRSV strategy.

Citations

Citations to this article as recorded by

Role of microRNAs in host defense against porcine reproductive and respiratory syndrome virus infection: a hidden front line
Xuewei Huang, Weiye Liu
Frontiers in Immunology.2024;[Epub] CrossRef
Porcine reproductive and respiratory syndrome virus infection induces microRNA novel-216 production to facilitate viral-replication by targeting MAVS 3´UTR
Xuegang Luo, Sha Xie, Xingsheng Xu, Yao Zhang, Yun Huang, Dongmei Tan, Yi Tan
Veterinary Microbiology.2024; 292: 110061. CrossRef
Antiviral Strategies against PRRSV Infection
Taofeng Du, Yuchen Nan, Shuqi Xiao, Qin Zhao, En-Min Zhou
Trends in Microbiology.2017; 25(12): 968. CrossRef
Anti-PRRSV effect and mechanism of tetrahydroaltersolanol Cin vitro
Song-Lin Zhang, Yi-Chun Wu, Fan Cheng, Zhi-Yong Guo, Jian-Feng Chen
Journal of Asian Natural Products Research.2016; 18(3): 303. CrossRef
Cellular microRNA miR-26a suppresses replication of porcine reproductive and respiratory syndrome virus by activating innate antiviral immunity
Xiaojuan Jia, Yuhai Bi, Jing Li, Qing Xie, Hanchun Yang, Wenjun Liu
Scientific Reports.2015;[Epub] CrossRef
Inhibition of porcine reproductive and respiratory syndrome virus replication in vitro using DNA-based short antisense oligonucleotides
Longlong Zheng, Xiang Li, Lingyun Zhu, Wengui Li, Junlong Bi, Guishu Yang, Gefen Yin, Jianping Liu
BMC Veterinary Research.2015;[Epub] CrossRef

Research Support, Non-U.S. Gov't

Biochemical Characteristics of Immune-Associated Phospholipase A2 and Its Inhibition by an Entomopathogenic Bacterium, Xenorhabdus nematophila: Sony Shrestha , Yonggyun Kim; J. Microbiol. 2009;47(6):774-782. Published online February 4, 2010; DOI: https://doi.org/10.1007/s12275-009-0145-3

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Abstract: An entomopathogenic bacterium, Xenorhabdus nematophila, induces an immunosuppression of target insects by inhibiting phospholipase A2 (PLA2) activity. Recently, an immune-associated PLA2 gene was identified from the red flour beetle, Tribolium castaneum. This study cloned this PLA2 gene in a bacterial expression vector to produce a recombinant enzyme. The recombinant T. castaneum PLA2 (TcPLA2) exhibited its characteristic enzyme activity with substrate concentration, pH, and ambient temperature. Its biochemical characteristics matched to a secretory type of PLA2 (sPLA2) because its activity was inhibited by dithiothreitol (a reducing agent of disulfide bond) and bromophenacyl bromide (a specific sPLA2 inhibitor) but not by methylarachidonyl fluorophosphonate (a specific cytosolic type of PLA2). The X. nematophila culture broth contained PLA2 inhibitory factor(s), which was most abundant in the media obtained at a stationary bacterial growth phase. The PLA2 inhibitory factor(s) was heat-resistant and extracted in both aqueous and organic fractions. Effect of a PLA2-inhibitory fraction on the immunosuppression of T. castaneum was equally comparable with that resulted from inhibition of the TcPLA2 gene expression by RNA interference.

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