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کاربرد نوع شرط:
- جایگاه : پژوهشی
- مجله: International Journal of Molecular and Clinical Microbiology
- نوع مقاله: Journal Article
- کلمات کلیدی: Persian Gulf,Cloning,Protease,Streptomyces,E. coli Xl1 blue
- چکیده:
- چکیده انگلیسی: Protease is an enzyme with various uses in medicine, industry and textile. One of the most important sources of protease production is bacteria such as Streptomyces. So, the aim of this study was cloning and sequencing of the protease gene in the Streptomyces spp isolated from Persian Gulf in Escherichia coli XL1blue. After collection of marine sediments from the Persian Gul, Streptomyces strains were identified using standard laboratory tests. All isolates were confirmed using 16S rRNA amplification test. Protease encoded gene were identified using specific primers in the PCR method. Protease gene was cloned in the E. coli host vector by TA cloning technique and finally the expression of the genes was measured using Real-time PCR method. ClustalX and Mega5 software were used to draw the phylogenetic tree. Twelve isolates of Streptomyces were isolated and 25% (n; 3/12) of them were positive for protease gene. After cloning of the gene, colony selection (blue / white colonies) were used for identification of success cloned strains. A relative expression of the protease gene was shown by real-time PCR test. Phylogenetic tree with the neighbor joining method show that, Streptomyces spp with bootstrap values 99% located in a clade which indicated their close relatedness. Protease enzyme production was performed by recombinant plasmid and TA cloning, and further studies could be helpful to optimize different conditions for this enzyme production. So, The Persian Gulf is a large pool for the protease producing Streptomyces for medical and industrial use.
- انتشار مقاله: 30-03-1399
- نویسندگان: Farnaz Heydari,Elahe Aliasgari,Kumarss Amini
- مشاهده
- جایگاه : پژوهشی
- مجله: International Journal of Molecular and Clinical Microbiology
- نوع مقاله: Journal Article
- کلمات کلیدی: aspirin,Real-time PCR,Fluconazole,Candida glabrata,ERG11
- چکیده:
- چکیده انگلیسی: Azole compounds have been a treatment option for Candida glabrata infections. However, azole resistance can occur through different mechanisms such as alterations in ERG11 (lanosterol 14α-demethylase). Aspirin (ASA), a non-steroidal anti-inflammatory drug, showed antimicrobial activity against Candida. The purpose of this study was to evaluate the synergistic effect of ASA as an anti-inflammatory drug with the fluconazole on the azole-resistant C. glabrata isolates. In the cross-sectional study, a total of 60 oral samples were collected from the school of dentistry, Tehran University of medical sciences. After confirmation of fungal isolates, template DNA was extracted and PCR for detection of Erg11 gene by specific primers was performed in all Fluconazole-resistant isolates. MIC for ASA was determined using broth dilution method in 96‐well plates. RNA was extracted and cDNA synthesized according to the Omniscript RT kit instructions. The effect of ASA in the treated and non-treated groups on ERG11 gene expression was determined by Real-Time PCR technique. Out of 60 collected samples, 12 (20%) C. glabrata were isolated. All of these isolates were resistant to fluconazole and carried ERG11 genes. Real-time PCR results showed that the combination of ASA with fluconazole reduced ERG11 gene expression. It is concluded that, treatment of candidal infections with ASA significantly reduced resistance in to azole compounds by down expression of ERG11 gene, suggesting that an NSAID might be useful for azole-resistance candidal infections.
- انتشار مقاله: 14-01-1398
- نویسندگان: Zahra Khosravi,Kumarss Amini,Mansour Bayat,Laya Takbiri Osgoei
- مشاهده