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Kui Wu,Yangyun Zheng,Qingping Wu,Haiying Chen,Songzhe Fu,Biao Kan,Yongyan Long,Xiansheng Ni,Junling Tu 한국미생물학회 2019 The journal of microbiology Vol.57 No.12
In order to adapt to different environments, Vibrio parahaemolyticus employed a complicated quorum sensing system to orchestrate gene expression and diverse colony morphology patterns. In this study, the function of the putative quorum sensing signal synthase gene cqsA (VPA0711 in V. parahaemolyticus strain RIMD2210633 genome) was investigated. The cloning and expression of V. parahaemolyticus cqsA in Escherichia coli system induced the production of a new quorum sensing signal that was found in its culture supernatant. The signal was purified by high performance liquid chromatography methods and determined to be 3-hydroxyundecan- 4-one by indirect and direct mass spectra assays. The deletion of cqsA in RIMD2210633 changed V. parahaemolyticus colony morphology from the classical ‘fried-egg’ shape (thick and opaque in the center, while thin and translucent in the edge) of the wild-type colony to a ‘pancake’ shape (no significant difference between the centre and the edge) of the cqsAdeleted colony. This morphological change could be restored by complementary experiment with cqsA gene or the signal extract. In addition, the expression of opaR, a well-known quorum sensing regulatory gene, could be up-regulated by cqsA deletion. Our results suggested that V. parahaemolyticus used cqsA to produce 3-hydroxyundecan-4-one signal and thereby regulated colony morphology and other quorum sensing-associated behaviors.
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Guangdong Hu,Jing Wang,Hui Huang,Fusheng Quan,Jian Kang,Yongyan Wu,Yuanpeng Gao,Feng Su,Minghao Shao,Yong Zhang 한국생물공학회 2014 Biotechnology and Bioprocess Engineering Vol.19 No.2
Transposons are widely used for geneticengineering in various model organisms. Recently, piggyBac(PB) has been developed as a transposable and efficientgene transfer tool in mammalian cells. In the present study,we developed three types of PB transposon systemscontaining a dual plasmid system (DPS), a single plasmidsystem (SPS), and a DNA-mRNA combined system (DRPS)and characterized their basic properties in HEK293 cells. The basic elements of the donor plasmid included aselectable-reporter gene expression cassette, two loxP sitesin the same orientation, a multiple cloning site, and twochicken β-globin insulator core elements. We further identifiedthe function of the selectable-reporter and examined PBintegration sites in the human genome. Moreover, wecompared the transposition efficacy and found that SPStransposed more efficiently, as compared to DPS; integrationinto the host genome was determined by measuring PBaseactivity. Results discovered the loss of PBase activity in theDRPS, indicating that this system is much more biologicallysafe, as compared to DPS and SPS. Finally, we employedthe DRPS to successfully perform a gene delivery intobovine mammary epithelial cells (BMECs). Taken together,the information from this study will improve the flexibilityof PB transposon systems and reduce the genotoxicity ofPBase in genetic engineering.
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