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  1. Wong CF, Rahman RNZRA, Basri M, Salleh AB
    Iran J Biotechnol, 2017;15(3):194-200.
    PMID: 29845069 DOI: 10.15171/ijb.1524
    Background:Pseudomonas protein expression in E. coli is known to be a setback due to significant genetic variation and absence of several genetic elements in E. coli for regulation and activation of Pseudomonas proteins. Modifications in promoter/repressor system and shuttle plasmid maintenance have made the expression of stable and active Pseudomonas protein possible in both Pseudomonas sp. and E. coli. Objectives: Construction of shuttle expression vectors for regulation and overexpression of Pseudomonas proteins in Pseudomonas sp. and E. coli. Materials and Methods:Pseudomonas-Escherichia shuttle expression vectors, pCon2(3), pCon2(3)-Kan and pCon2(3)-Zeo as well as E. coli expression vectors of pCon4 and pCon5 were constructed from pUCP19-, pSS213-, pSTBlue-1- and pPICZαA-based vectors. Protein overexpression was measured using elastase strain K as passenger enzyme in elastinolytic activity assay. Results: The integration of two series of IPTG inducible expression cassettes in pCon2(3), pCon2(3)-Kan and pCon2(3)-Zeo, each carrying an E. coli lac-operon based promoter, Plac, and a tightly regulated T7(A1/O4/O3) promoter/repressor system was performed to facilitate overexpression study of the organic solvent-tolerant elastase strain K. These constructs have demonstrated an elastinolytic fold of as high as 1464.4 % in comparison to other published constructs. pCon4 and pCon5, on the other hand, are series of pCon2(3)-derived vectors harboring expression cassettes controlled by PT7(A1/O4/O3) promoter, which conferred tight regulation and repression of basal expression due to existence of respective double operator sites, O3 and O4, and lacIq. Conclusions: The constructs offered remarkable assistance for overexpression of heterogeneous genes in Pseudomonas sp. and E. coli for downstream applications such as in industries and structural biology study.
  2. Hossain K, Quaik S, Ismail N, Rafatullah M, Avasan M, Shaik R
    Iran J Biotechnol, 2016 Sep;14(3):154-162.
    PMID: 28959331 DOI: 10.15171/ijb.1216
    BACKGROUND: Application of membrane technology to wastewater treatment has expanded over the last decades due to increasingly stringent legislation, greater opportunities for water reuse/recycling processes and continuing advancement in membrane technology.

    OBJECTIVES: In the present study, a bench-scale submerged microfiltration membrane bioreactor (MBR) was used to assess the treatment of textile wastewater.

    MATERIALS AND METHODS: The decolorization capacity of white-rot fungus coriolus versicolor was confirmed through agar plate and liquid batch studies. The temperature and pH of the reactor were controlled at 29±1°C and 4.5±2, respectively. The bioreactor was operated with an average flux of 0.05 m.d(-1) (HRT=15hrs) for a month.

    RESULTS: Extensive growth of fungi and their attachment to the membrane led to its fouling and associated increase of the transmembrane pressure requiring a periodic withdrawal of sludge and membrane cleaning. However, stable decoloration activity (approx. 98%), BOD (40-50%), COD (50-67%) and total organic carbon (TOC) removal (>95%) was achieved using the entire system (fungi + membrane), while the contribution of the fungi culture alone for TOC removal, as indicated by the quality of the reactor supernatant, was 35-50% and 70%, respectively.

    CONCLUSIONS: The treated wastewater quality satisfied the requirement of water quality for dyeing and finishing process excluding light coloration. Therefore, textile wastewater reclamation and reuse is a promising alternative, which can both conserve or supplement the available water resource and reduce or eliminate the environmental pollution.

  3. Elyasi Gorji Z, Amiri-Yekta A, Gourabi H, Hassani S, Fatemi N, Zerehdaran S, et al.
    Iran J Biotechnol, 2015 Jun;13(2):10-17.
    PMID: 28959285 DOI: 10.15171/ijb.1004
    BACKGROUND: Follicle stimulating hormone (FSH) plays an essential role in reproductive physiology and follicular development.

    OBJECTIVE: A new variant of the equine fsh (efsh) gene was cloned, sequenced, and expressed in Pichia pastoris (P. pastoris) GS115 yeast expression system.

    MATERIALS AND METHODS: The full-length cDNAs of the efshα and efshβ chains were amplified by reverse transcription polymerase chain reaction (RT-PCR) using the total RNA isolated from an Iranian Turkmen-thoroughbred horse's anterior pituitary gland. The amplified efsh chains were cloned into the pPIC9 vector and transferred into P. pastoris. The secretion of recombined eFSH using P. pastoris expression system was confirmed by Western blotting and immunoprecipitation (IP) methods.

    RESULTS: The DNA sequence of the efshβ chain accession number JX861871, predicted two putative differential nucleotide arrays, both of which are located in the 3'UTR. Western blotting showed a molecular mass of 13 and 18 kDa for eFSHα and eFSHβ subunits, respectively. The expression of desired protein was confirmed by protein G immunoprecipitation kit.

    CONCLUSIONS: eFSH successfully expressed in P. pastoris. These findings lay a foundation to improve ovulation and embryo recovery rates as well as the efficiency of total embryo-transfer process in mares.

  4. Kannan M, Mohamad Saad M, Zainal Z, Kassim H, Ismail I, Talip N, et al.
    Iran J Biotechnol, 2020 Oct;18(4):e2566.
    PMID: 34056024 DOI: 10.30498/IJB.2020.2566
    Background: Rice tungro disease (RTD) is a viral disease mainly affecting rice in Asia. RTD caused by Rice tungro bacilliform virus and Rice tungro spherical virus. To date, there are only 5 RTSV isolates have been reported.

    Objectives: In this study, we aimed to report the complete nucleotide sequence of Malaysian isolate of Rice tungro spherical virus Seberang Perai (RTSV-SP) for the first time. RTSV-SP was characterized and its evolutionary relationship with previously reported Indian and Philippines isolates were elucidated.

    Materials and Methods: RTSV-SP isolate was isolated from a recent outbreak in a paddy field in Seberang Perai zone of Malaysia. Its complete genome was amplified by RT-PCR, cloned and sequenced.

    Results: Sequence analysis indicated that the genome of RTSV-SP consisted of 12,173 nucleotides (nt). Comparative analysis of 6 complete genome sequences using Clustal Omega showed that Seberang Perai isolate shared the highest nucleotide identity (96.04%) with Philippine-A isolate, except that the sORF-2 of RTSV-SP is shorter than RTSV Philippine-A by 27 amino acid residues. RTSV-SP found to cluster in Southeast Asia (SEA) group based on the whole genome sequence phylogenetic analysis using MEGA X software.

    Conclusions: Phylogenetic classification of RTSV isolates based on the complete nucleotide sequences showed more distinctive clustering pattern with the addition of RTSV-SP whole genome to the available isolates. Present study described the isolation and molecular characterization of RTSV-SP.

  5. A Al-Kafaween M, Mohd Hilmi AB, A Nagi Al-Jamal H, A Elsahoryi N, Jaffar N, Khairi Zahri M
    Iran J Biotechnol, 2020 Oct;18(4):e2542.
    PMID: 34056021 DOI: 10.30498/IJB.2020.2542
    Background: Honey has been known as a traditional medicine for centuries with its antibacterial properties. It is considered one of the most enduring substances used in wound management.

    Objectives: This study aimed to: (i) evaluate the effects of Malaysian Trigona honey on bacterial structure and (ii) assess the anti-virulence potential of this honey by examining their impacts on the expression of selected genes (involved in stress survival and biofilm formation) in a test organism.

    Materials and Methods: Trigona honey's impacts on the bacterial structure (cell morphology) and the expression profiles of select Pseudomonas Aeruginosa and Streptococcus Pyogenes genes were examined using scanning electron microscopy (SEM) and real-time PCR (RT-qPCR) analysis, respectively.

    Results: SEM showed that the decreased cell density deformed, disrupted, and damaged cells for both bacteria. RT-qPCR showed that the expression of fleN, fleQ, and fleR genes of P.aeruginosa were decreased, 4.26-fold, 3.80-fold and 2.66- fold respectively. In addition, scpA, ftsY, and emm13 of S.pyogenes were decreased, 2.87-fold, 3.24-fold, and 4.65-fold respectively.

    Conclusion: Our results indicate that Trigona honey may be an effective inhibitor and virulence modulator of P. aeruginosa and S. pyogenes via multiple molecular targets. This deduction needs to be investigated in vivo.

  6. Tavakoli A, Hamzah A
    Iran J Biotechnol, 2017 Mar;15(1):67-73.
    PMID: 28959354 DOI: 10.15171/ijb.1344
    BACKGROUND: Benzaldehyde dehydrogenase (BZDH) is encoded by the xylC that catalyzes the conversion of benzaldehyde into benzoate in many pathways such as toluene degradation.

    OBJECTIVES: In this study, the xylC gene from Rhodococcus ruber UKMP-5M was expressed in Escherichia coli, purified, and characterized.

    MATERIALS AND METHODS: The xylC was amplified and cloned in E. coli. The recombinant plasmid pGEMT-xylC was digested by NdeI and HindIII to construct plasmid pET28b-xylC and transformed in E. coli BL21 (DE3). Expression of the recombinant protein was induced by 1 mM isopropyl β-D-thiogalactoside (IPTG) at 37°C. The BZDH was purified by ion exchange chromatography, in which the product was an NAD-dependent enzyme using benzaldehyde as a substrate for enzyme characterization. The end metabolite was identified via gas chromatography mass spectrometry (GC-MS).

    RESULTS: The recombinant BZDH is 27 kDa, purified by ion exchange chromatography. The activity of BZDH was 9.4 U.μL-1 The optimum pH and temperature were 8.5 and 25ºC, respectively. The Michaelis constant (Km) and maximum velocity (Vmax) were 4.2 mM and 19.7 U.mL-1, respectively. The metabolite of BZDH was benzene carboxylic acid as determined by GC-MS analysis.

    CONCLUSIONS: BZDH has the ability to degrade benzaldehyde to less toxic compounds. The BZDH is a critical enzyme for the degradation of aromatic hydrocarbons in Rhodococcus sp. The BZDH from R. ruber UKMP-5M is showed similar function with other aldehyde dehydrogenases.

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