Browsing by Author "Chang, Sheng-Nan"

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Effective and site-specific phosphoramidation reaction for universally labeling nucleic acids
(2014-03-15)
Su, Yu-Chih
;
Chen, Hsing-Yin
;
Ko, Ni Chien
;
Hwang, Chi-Ching
;
Wu, Min Hui
Here we report efficient and selective postsynthesis labeling strategies, based on an advanced phosphoramidation reaction, for nucleic acids of either synthetic or enzyme-catalyzed origin. The reactions provided phosphorimidazolide intermediates of DNA or RNA which, whether reacted in one pot (one-step) or purified (two-step), were directly or indirectly phosphoramidated with label molecules. The acquired fluorophore-labeled nucleic acids, prepared from the phosphoramidation reactions, demonstrated labeling efficacy by their F/N ratio values (number of fluorophores per molecule of nucleic acid) of 0.02-1.2 which are comparable or better than conventional postsynthesis fluorescent labeling methods for DNA and RNA. Yet, PCR and UV melting studies of the one-step phosphoramidation-prepared FITC-labeled DNA indicated that the reaction might facilitate nonspecific hybridization in nucleic acids. Intrinsic hybridization specificity of nucleic acids was, however, conserved in the two-step phosphoramidation reaction. The reaction of site-specific labeling nucleic acids at the 5'-end was supported by fluorescence quenching and UV melting studies of fluorophore-labeled DNA. The two-step phosphoramidation-based, effective, and site-specific labeling method has the potential to expedite critical research including visualization, quantification, structural determination, localization, and distribution of nucleic acids in vivo and in vitro. (C) 2013 Elsevier Inc. All rights reserved.
Successful preparation and characterization of biotechnological grade agarose from indigenous Gelidium amansii of Taiwan
(2012-03-01)
Wang, Tzu-Pin
;
Chang, Li-Lin
;
Chang, Sheng-Nan
;
Wang, Eng-Chi
;
Hwang, Long-Chih
This paper reports the first successful preparation of biotechnological grade agarose from Gelidium amansii found in Taiwan. The scale-efficiency preparation was achieved by shortening EDTA treatment time through dispersing G. amansii agar in water in the presence of heat and EDTA, removing agaropectin impurity with a heat-compatible and strong-anion exchange resin, and precipitating agarose with a cost-effective isopropanol method. The yield of agarose from prepared G. amansii agar was 11.3%. The acquired agarose has a gel strength of 853 g cm(-2), a sulfate content of 0.14%, a pyruvate content of 1.03%, a degree of electroendosmosis of 0.16 and very limited binding affinity to DNA. The excellent properties of agarose from G. amansii of Taiwan confirm its potential diverse biotechnological applications. This innovative agarose preparation method with the significantly improved scale-efficiency can be modified for large-scale preparation of agarose for use in biotechnological industry and biochemical research. (C) 2011 Elsevier Ltd. All rights reserved.