Bacterial Luciferase Gene Cassette as a Real-time Bioreporter for Infection Model and Drug Evaluation

2018 ◽  
Vol 24 (8) ◽  
pp. 952-958 ◽  
Author(s):  
Xiwen Wang ◽  
Hang Chi ◽  
Bo Zhou ◽  
Wenliang Li ◽  
Zhiping Li ◽  
...  
Keyword(s):  
Real Time ◽  
Cell Lines ◽  
Drug Evaluation ◽  
Gene Cassette ◽  
Infection Model ◽  
High Signal ◽  
Luciferase Gene ◽  
Bacterial Luciferase ◽  
Lux Operon ◽  
Wide Range

The bacterial luciferase gene cassette (lux) is an ideal bioreporter for real-time monitoring of the dynamics of bacteria because it is a fully autonomous, substrate-free bioluminescent reporter system available in a prokaryotic or eukaryotic host background. The lux operon is emerging as a powerful bioreporter for the study of a wide range of biological processes such as gene function, drug discovery and development, cellular trafficking, protein-protein interactions, and especially tumorigenesis and cancer treatment. Furthermore, the use of a high signal to noise bioluminescent bioreporter is quickly replacing traditional fluorescent bioreporter because of the lack of endogenous bioluminescent reactions in living animals. This review briefly describes how the lux operon is used for bioluminescence imaging. Current advances in bioluminescence bacteria development are summarized, focusing on their construction strategy and applications in bacterial infection and antibiotic treatment. Different construction methods of lux-expressing cell lines are also discussed. Taken together, this review provides valuable guidelines toward the development of an ideal bioluminescent bacteria or cell lines to evaluate the efficacy of a drug.

scholarly journals The Evolution of the Bacterial Luciferase Gene Cassette (lux) as a Real-Time Bioreporter

Sensors ◽  
2012 ◽  
Vol 12 (1) ◽  
pp. 732-752 ◽  
Author(s):  
Dan Close ◽  
Tingting Xu ◽  
Abby Smartt ◽  
Alexandra Rogers ◽  
Robert Crossley ◽  
...  
Keyword(s):  
Real Time ◽  
Gene Cassette ◽  
Luciferase Gene ◽  
Bacterial Luciferase

scholarly journals Autonomous Bioluminescent Expression of the Bacterial Luciferase Gene Cassette (lux) in a Mammalian Cell Line

PLoS ONE ◽  
2010 ◽  
Vol 5 (8) ◽  
pp. e12441 ◽  
Author(s):  
Dan M. Close ◽  
Stacey S. Patterson ◽  
Steven Ripp ◽  
Seung J. Baek ◽  
John Sanseverino ◽  
...  
Keyword(s):  
Cell Line ◽  
Mammalian Cell ◽  
Gene Cassette ◽  
Mammalian Cell Line ◽  
Luciferase Gene ◽  
Bacterial Luciferase

scholarly journals Introducing Novel Molecular-based Method for Quantification of Homologous Recombination Efficiency

2021 ◽  
Author(s):  
Mustapha Dibbasey ◽  
Terry Gaymes
Keyword(s):  
Homologous Recombination ◽  
Real Time ◽  
Cell Lines ◽  
Real Time Pcr ◽  
Cost Effective ◽  
Lacz Gene ◽  
Expression Arrays ◽  
Recombinant Product ◽  
Wide Range ◽  
High Level

AbstractBackgroundHomologous recombination (HR) pathway is a DNA double-stranded breaks repair pathway well-known for its high level of accuracy. Low HR pathway efficiency clinically known as homologous recombination deficiency (HRD) was identified in some cancers such as breast and ovarian cancers and studies have reported the sensitivity of HRD cancer cells to DNA repair inhibitors such as Olaparib. However, current techniques including immunofluorescence-based technique are qualitative-based, hence lack sensitivity to determine the functionality of HR pathway. Additionally, some of the techniques including gene expression arrays require expression study of wide range genes involve in HR pathway, which is not cost-effective. The aim of the study is to optimise a PCR-based assay (Norgen’s Homologous Recombination kit) that can be employed to quantitate HR efficiency in cells, which accurately reflects the functional status of HR pathway.Methods and FindingsThe kit has two test plasmids (dl-1 and dl-2) with partial deletions in the LacZ gene and the plasmids are generated from modification of pUC19. HR-proficient (HeLa and AsPC-1) and HR-deficient (CAPAN-1 cells) cancer cell lines were transfected with the two plasmids to generate functional LacZ gene (i.e. recombinant product). The recombinant product was quantified by real-time PCR. Although recombinant product was generated in all the cell lines, our real-time PCR demonstrated a high quantity of recombinant product in HeLa cell line whilst low quantity in CAPAN-1 and AsPC-1 cell lines. The quantity of recombinant product generated and quantified reflects HR pathway efficiency.ConclusionOverall, the results have provided some evidence that the PCR-based kit can be suitably employed for quantification of HR efficiency provided appropriate transfection method and reagent are used. However, further study is required to confirm HR efficiency status of AsPC-1 cells to ascertain the low HR efficiency detected by the kit in these cells.

Light without substrate amendment: the bacterial luciferase gene cassette as a mammalian bioreporter

2011 ◽  
Author(s):  
Dan M. Close ◽  
Tingting Xu ◽  
Abby E. Smartt ◽  
Pat Jegier ◽  
Steven A. Ripp ◽  
...  
Keyword(s):  
Gene Cassette ◽  
Luciferase Gene ◽  
Bacterial Luciferase ◽  
Substrate Amendment

Phyto-Facilitated Bimetallic ZnFe2O4 Nanoparticles via Boswellia carteri: Synthesis, Characterization and Anti-Cancer Activity.

2020 ◽  
Vol 21 ◽  
Author(s):  
Amer Imraish ◽  
Afnan Al-Hunaiti ◽  
Tuqa Abu-Thiab ◽  
Abed Al-Qader Ibrahim ◽  
Eman Hwaitat ◽  
...  
Keyword(s):  
Anticancer Activity ◽  
Cell Lines ◽  
Metallic Nanoparticles ◽  
Bimetallic Nanoparticles ◽  
Iron Chloride ◽  
Nano Particles ◽  
Adverse Side Effect ◽  
Wide Range ◽  
Anti Cancer ◽  
Znfe2o4 Nanoparticles

Background: The growing unsatisfaction toward the available traditional chemotherapeutic agents enhanced the need to develop new methods for obtaining materials with more effective and safe anti-cancer properties. Over the past few years, usage of metallic nanoparticles has been a target for researchers of different scientific and commercial fields due to their tiny sizes, environment friendly properties and wide range applications. To overcome the obstacles of traditional physical and chemical methods for synthesis of such nanoparticles, a new less expensive and eco-friendly method has been adopted using natural existing organisms as a reducing agent to mediate synthesis of the desired metallic nanoparticles from their precursors, a process called green biosynthesis of nanoparticles. Objective: Here in the present study, zinc iron bimetallic nanoparticles (ZnFe2O4) were synthesized via an aqueous extract of Boswellia Carteri resin mixed with zinc acetate and iron chloride precursors, and they were tested for their anticancer activity. Methods: Various analytic methods were applied for the characterization of the Phyto synthesized ZnFe2O4 and they were tested for their anticancer activity against MDA-MB-231, K562, MCF-7 cancer cell lines and normal fibroblasts. Results: Our results demonstrate the synthesis of cubic structured bimetallic nanoparticles ZnFe2O4 with an average diameter 10.54 nm. MTT cytotoxicity assay demonstrate that our phyto-synthesized ZnFe2O4 nanoparticles exhibited a selective and potent anticancer activity against K562 and MDA-MB-231 cell lines with IC50 values 4.53 µM and 4.19 µM, respectively. Conclusion: In conclusion, our bio synthesized ZnFe2O4 nano particles show a promising environmentally friendly of low coast chemotherapeutic approach against selective cancers with a predicted low adverse side effect toward normal cells. Further in vivo advanced animal research should be done to execute their applicability in living organisms.

Synthesis and Biological Evaluation of Novel 4,5,6,7-Tetrahydrobenzo[D]-Thiazol-2- Yl Derivatives Derived from Dimedone with Anti-Tumor, C-Met, Tyrosine Kinase and Pim-1 Inhibitions

2019 ◽  
Vol 19 (12) ◽  
pp. 1438-1453 ◽  
Author(s):  
Rafat M. Mohareb ◽  
Amr S. Abouzied ◽  
Nermeen S. Abbas
Keyword(s):  
Tyrosine Kinase ◽  
Tumor Cell ◽  
Cell Lines ◽  
Single Molecule ◽  
Anticancer Agents ◽  
Biological Evaluation ◽  
New Drugs ◽  
Tumor Cell Lines ◽  
Thiazole Derivatives ◽  
Wide Range

Background: Dimedone and thiazole moieties are privileged scaffolds (acting as primary pharmacophores) in many compounds that are useful to treat several diseases, mainly tropical infectious diseases. Thiazole derivatives are a very important class of compounds due to their wide range of pharmaceutical and therapeutic activities. On the other hand, dimedone is used to synthesize many therapeutically active compounds. Therefore, the combination of both moieties through a single molecule to produce heterocyclic compounds will produce excellent anticancer agents. Objective: The present work reports the synthesis of 47 new substances belonging to two classes of compounds: Dimedone and thiazoles, with the purpose of developing new drugs that present high specificity for tumor cells and low toxicity to the organism. To achieve this goal, our strategy was to synthesize a series of 4,5,6,7-tetrahydrobenzo[d]-thiazol-2-yl derivatives using the reaction of the 2-bromodimedone with cyanothioacetamide. Methods: The reaction of 2-bromodimedone with cyanothioacetamide gave the 4,5,6,7-tetrahydrobenzo[d]- thiazol-2-yl derivative 4. The reactivity of compound 4 towards some chemical reagents was observed to produce different heterocyclic derivatives. Results: A cytotoxic screening was performed to evaluate the performance of the new derivatives in six tumor cell lines. Thirteen compounds were shown to be promising toward the tumor cell lines which were further evaluated toward five tyrosine kinases. Conclusion: The results of antitumor screening showed that many of the tested compounds were of high inhibition towards the tested cell lines. Compounds 6c, 8c, 11b, 11d, 13b, 14b, 15c, 15g, 21b, 21c, 20d and 21d were the most potent compounds toward c-Met kinase and PC-3 cell line. The most promising compounds 6c, 8c, 11b, 11d, 13b, 14b, 15c, 15g, 20c, 20d, 21b, 21c and 21d were further investigated against tyrosine kinase (c-Kit, Flt-3, VEGFR-2, EGFR, and PDGFR). Compounds 6c, 11b, 11d, 14b, 15c, and 20d were selected to examine their Pim-1 kinase inhibition activity the results revealed that compounds 11b, 11d and 15c had high activities.

Heterocyclization of 2-(2-phenylhydrazono)cyclohexane-1,3-dione to Synthesis Thiophene, Pyrazole and 1,2,4-triazine Derivatives with Anti-Tumor and Tyrosine Kinase Inhibitions

2020 ◽  
Vol 20 (10) ◽  
pp. 1209-1220
Author(s):  
Rafat M. Mohareb ◽  
Ensaf S. Alwan
Keyword(s):  
Cell Lines ◽  
Cancer Cell ◽  
Tyrosine Kinases ◽  
Proliferative Activity ◽  
Cancer Cell Lines ◽  
Thiazole Derivatives ◽  
Wide Range ◽  
Cytotoxic Compounds ◽  
Triazine Derivatives ◽  
Target Molecules

Background: Recently tetrahydrobenzo[b]thiazole derivatives acquired a special attention due to their wide range of pharmacological activities especially the therapeutic activities. Through the market it was found that many pharmacological drugs containing the thiazole nucleus were known. Objective: This work aimed to synthesize target molecules not only possess anti-tumor activities but also kinase inhibitors. The target molecules were obtained starting from the arylhydrazonocyclohexan-1,3-dione followed by their heterocyclization reactions to produce anticancer target molecules. Methods: The arylhydrazone derivatives 3a-c underwent different heterocyclization reactions to produce thiophene, thiazole, pyrazole and 1,2,4-triazine derivatives. The anti-proliferative activity of twenty six compounds among the synthesized compounds toward the six cancer cell lines namely A549, H460, HT-29, MKN-45, U87MG, and SMMC-7721 was studied. Results: Anti-proliferative evaluations, tyrosine and Pim-1 kinase inhibitions were perform for most of the synthesized compounds where the varieties of substituent through the aryl ring and the thiophene moiety afforded compounds with high activities. Conclusion: The compounds with high anti-proliferative activity towards the cancer cell lines showed that compounds 3b, 3c, 5e, 5f, 8c, 9c, 11c, 12c, 14e, 14f and 16c were the most cytotoxic compounds. Further tests of the latter compounds toward the five tyrosine kinases c-Kit, Flt-3, VEGFR-2, EGFR, and PDGFR and Pim-1 kinase showed that compounds 3c, 5e, 5f, 8c, 9c, 12c, 14e, 14f and 16c were the most potent of the tested compounds toward the five tyrosine kinases and compounds 6d, 11a, 20b and 21e were of the highest inhibitions towards Pim-1 kinase. Pan Assay Interference Compounds (PAINS) for the most cytotoxic compounds showed zero PAINS alert and can be used as lead compounds.

New Approaches for the Synthesis of Fused Thiophene, Pyrazole , Pyran and Pyridine Derivatives with Anti-Proliferative Together with c-Met Kinase and Prostate Cancer Cell Inhibitions

2020 ◽  
Vol 20 ◽  
Author(s):  
Rafat M. Mohareb ◽  
Yara R. Milad ◽  
Reem A. El-Ansary
Keyword(s):  
Prostate Cancer ◽  
Cell Lines ◽  
Cancer Cell ◽  
Prostate Cancer Cell ◽  
Aromatic Aldehydes ◽  
Cancer Cell Lines ◽  
Enzymatic Activities ◽  
Pyridine Derivatives ◽  
Wide Range ◽  
Target Molecules

Background:: Recently multi-component reactions producing pyran and pyridine derivatives acquired a special attention due to their wide range of pharmacological activities especially the therapeutic activities. Through the market it was found that many pharmacological drugs containing the pyran and pyridine nucleus were known. Objective:: We are aiming in this work to synthesize target molecules not only possess anti-tumor activities but also kinase inhibitors. The target molecules were obtained starting from cyclohexan-1,3-dione followed by its heterocyclization reactions to produce anticancer target molecules. Methods:: This work demonstrated multi-component reactions of cyclohexan-1,3-dione with aromatic aldehydes and diethylmalonate using triethylamine as a catalyst to give the 7,8-dihydro-4H-chromen-5(6H)-one derivatives 4a-c. The reaction of compounds 4a-c with either of hydrazine hydrate of phenylhydrazine gave the chromeno[2,3-c]pyrazole derivatives 5a-f, respectively. In addition, further heterocyclization reactions were adopted to give the chromeno[3,2-d]isoxazole, chromene-3-carboxamide derivatives. Moreover, the multi-component reaction of cyclohexan-1,3-dione (1) with either of aromatic aldehydes and diethylmalonate using a catalytic amount of ammonium acetate gave the 1,4,5,6,7,8-hexahydroquinoline derivatives 13a-c. The anti-proliferative activities of the synthesized compounds toward the six cancer cell lines namely A549, H460, HT-29, MKN-45, U87MG, and SMMC-7721 were studied. In addition the c-Met enzymatic activities and inhibition toward the prostate cancer cell PC-3 were measured. Results:: Anti-proliferative evaluations, c-Met enzymatic activities and inhibition toward the prostate cancer cell PC-3 were measured and the results obtained in most cases, indicated that the presence of electronegative Cl group through the molecule favour the inhibitions. Conclusion:: The compounds with high anti-proliferative activity towards the cancer cell lines were 4a, 4b, 6d, 6e, 6f, 10e, 10f, 12c, 14e, 14f, 15c, 16d, 16e, 16f, 19c and 20c. Compounds 4b, 6c, 6d, 8b, 10c, 10d, 12b, 13b, 14c, 14d, 15b, 16c, 16d, 17b, 17c, 19b, 20b and 20c exhibited high potency against c-Met kinase and compounds 4a, 4b, 6b, 6c, 6d, 6f, 8b, 8c, 10c, 10d, 10e, 12b, 12c, 13a, 13b, 13c, 14c, 14d, 14e, 14f, 15b, 15c, 16b, 16c, 16d, 17b, 17c, 19c, 19d, 20a, 20b and 20c displayed high inhibitions toward PC-3 cell line.

scholarly journals Development of Real-Time Time Gated Digital (TGD) OFDR Method and Its Performance Verification

Sensors ◽  
2021 ◽  
Vol 21 (14) ◽  
pp. 4865
Author(s):  
Kinzo Kishida ◽  
Artur Guzik ◽  
Ken’ichi Nishiguchi ◽  
Che-Hsien Li ◽  
Daiji Azuma ◽  
...  
Keyword(s):  
Real Time ◽  
Optical Fibers ◽  
High Speed ◽  
Oil And Gas ◽  
Scattered Light ◽  
Oil And Gas Industry ◽  
High Signal ◽  
Chirp Pulse ◽  
Sound Waves ◽  
Industry Applications

Distributed acoustic sensing (DAS) in optical fibers detect dynamic strains or sound waves by measuring the phase or amplitude changes of the scattered light. This contrasts with other distributed (and more conventional) methods, such as distributed temperature (DTS) or strain (DSS), which measure quasi-static physical quantities, such as intensity spectrum of the scattered light. DAS is attracting considerable attention as it complements the conventional distributed measurements. To implement DAS in commercial applications, it is necessary to ensure a sufficiently high signal-noise ratio (SNR) for scattered light detection, suppress its deterioration along the sensing fiber, achieve lower noise floor for weak signals and, moreover, perform high-speed processing within milliseconds (or sometimes even less). In this paper, we present a new, real-time DAS, realized by using the time gated digital-optical frequency domain reflectometry (TGD-OFDR) method, in which the chirp pulse is divided into overlapping bands and assembled after digital decoding. The developed prototype NBX-S4000 generates a chirp signal with a pulse duration of 2 μs and uses a frequency sweep of 100 MHz at a repeating frequency of up to 5 kHz. It allows one to detect sound waves at an 80 km fiber distance range with spatial resolution better than a theoretically calculated value of 2.8 m in real time. The developed prototype was tested in the field in various applications, from earthquake detection and submarine cable sensing to oil and gas industry applications. All obtained results confirmed effectiveness of the method and performance, surpassing, in conventional SM fiber, other commercially available interrogators.

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