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Rafique B.,Government College University at Faisalabad | Khalid A.M.,University of Sargodha | Akhtar K.,National Institute for Biotechnology and Genetic Engineering NIBGE | Jabbar A.,Government College University at Faisalabad
Biosensors and Bioelectronics | Year: 2013

Electrochemical DNA biosensor was used to study the interaction of methotrexate (MTX) with DNA immobilized on the bare surface of glassy carbon electrode (GCE). The binding mechanism of MTX with DNA was elucidated by using constant current potentiometric technique further supported by UV-Visible and FT-IR studies. The decrease in guanine peak area was used as an analytical signal for the interaction of drug with DNA in acetate buffer solution at pH 4.2 (20% ethanol). The binding constant (K) value calculated for MTX was 3.821×105M-1. UV-Visible studies indicated hyperchromic and hypsochromic shifts in the maximum absorption bands of MTX after interaction with DNA. FT-IR investigations of MTX-DNA interaction revealed significant changes in the characteristic IR absorption bands of all the bases and phosphate groups of DNA. Furthermore, the shift of characteristics bands of C=O, N-H, C-H and O-H groups of MTX endow evidence for the interaction of MTX with DNA supporting the intercalative binding between them. © 2012 Elsevier B.V. Source


Tariq A.,National Institute for Biotechnology and Genetic Engineering NIBGE
Canadian journal of microbiology | Year: 2012

Bacillary dysentery, common in developing countries, is usually caused by Shigella species. A major problem in shigellosis is the rapid emergence of multidrug-resistant strains. This is the first detailed molecular study on drug resistance of Shigella isolates from the Faisalabad region of Pakistan. Ninety-five Shigella isolates obtained after screening of 2500 stool samples were evaluated for in vitro resistance to commonly used antimicrobial agents; the presence or absence of 20 of the most relevant drug resistance genes; and the prevalence of integrons 1, 2, and 3. Shigella flexneri was found to be the most prevalent and most resistant species. Collectively, high resistance was found towards ampicillin (96.84%), tetracycline (93.68%), streptomycin (77.89%), and chloramphenicol (72.63%). Significant emerging resistance was detected towards the modern frontline drugs ciprofloxacin (12.63%), cefradine (17.89%), ceftriaxone (20.00%), cefoperazone (22.10%), and cefixime (28.42%). Prevalence rates for bla(TEM), bla(CTX-M), gyrA, gyrB, qnrS, aadA1, strAB, tetA, tetB, catA, and catP were 78.94%, 12.63%, 20.00%, 21.05%, 21.05%, 67.36%, 42.10%, 12.63%, 53.68%, 33.68%, and 25.26%, respectively. Class 2 integrons (42.10%) were more common in the local isolates. Simultaneous detection of class 1 and 2 integrons in some isolates and a rapidly emerging resistance to modern frontline drugs are the major findings of this study. Source


Saeed M.,National Institute for Biotechnology and Genetic Engineering NIBGE
Australasian Plant Disease Notes | Year: 2010

Tomato leaf curl virus (ToLCV) from Australia is a monopartite begomovirus which is naturally associated with a DNA satellite, a vestigial betasatellite. Cotton leaf curl disease is caused by a complex consisting of one or more begomoviruses (eight species have been identified so far) associated with a single DNA β satellite named as Cotton leaf curl Multan betasatellite (CLCuMB). ToLCV and CLCuMB caused mild symptoms in cotton plants 1821 days post-inoculation. The mild symptoms caused by ToLCV and CLCuMB in cotton plants began to diminish 6 weeks post-inoculation and completely disappeared 810 weeks post-inoculation, raising the possibility that ToLCV may lack some factor(s) essential for persistent systemic infection of cotton. © Australasian Plant Pathology ociety 2010. Source


Rivera-Gil P.,University of Marburg | Nazarenus M.,University of Marburg | Ashraf S.,University of Marburg | Ashraf S.,National Institute for Biotechnology and Genetic Engineering NIBGE | Parak W.J.,University of Marburg
Small | Year: 2012

The concept of a long-term sensor for ion changes in the lysosome is presented. The sensor is made by layer-by-layer assembly of oppositely charged polyelectrolytes around ion-sensitive fluorophores, in this case for protons. The sensor is spontaneously incorporated by cells and resides over days in the lysosome. Intracellular changes of the concentration of protons upon cellular stimulation with pH-active agents are monitored by read-out of the sensor fluorescence at real time. With help of this sensor concept it is demonstrated that the different agents used (Monensin, Chloroquine, Bafilomycin A1, Amiloride) possessed different kinetics and mechanisms of action in affecting the intracellular pH values. The concept of a long-term sensor for ion changes in the lysosome is presented. The sensor is made by layer-by-layer assembly of oppositely charged polyelectrolytes around ion-sensitive fluorophores, in this case for protons. The sensor is spontaneously incorporated by cells and resides over days in the lysosome. Intracellular changes of the concentration of protons upon cellular stimulation with pH-active agents are monitored by read-out of the sensor fluorescence at real time. With help of this sensor concept it is demonstrated that the different agents used possess different kinetics and mechanisms of action in affecting the intracellular pH values. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Source


Ahmad N.,Imperial College London | Ahmad N.,National Institute for Biotechnology and Genetic Engineering NIBGE | Michoux F.,Imperial College London | Nixon P.J.,Imperial College London
PLoS ONE | Year: 2012

Chloroplast transformation provides an inexpensive, easily scalable production platform for expression of recombinant proteins in plants. However, this technology has been largely limited to the production of soluble proteins. Here we have tested the ability of tobacco chloroplasts to express a membrane protein, namely plastid terminal oxidase 1 from the green alga Chlamydomonas reinhardtii (Cr-PTOX1), which is predicted to function as a plastoquinol oxidase. A homoplastomic plant containing a codon-optimised version of the nuclear gene encoding PTOX1, driven by the 16S rRNA promoter and 5′UTR of gene 10 from phage T7, was generated using a particle delivery system. Accumulation of Cr-PTOX1 was shown by immunoblotting and expression in an enzymatically active form was confirmed by using chlorophyll fluorescence to measure changes in the redox state of the plastoquinone pool in leaves. Growth of Cr-PTOX1 expressing plants was, however, more sensitive to high light than WT. Overall our results confirm the feasibility of using plastid transformation as a means of expressing foreign membrane proteins in the chloroplast. © 2012 Ahmad et al. Source

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