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Capillary enzymophoresis of nucleic acid fragments using coupled capillary electrophoresis and capillary enzyme microreactors having surface‐immobilized RNA‐modifying enzymes
Author(s) -
Mechref Yehia,
El Rassi Ziad
Publication year - 1995
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.11501601349
Subject(s) - capillary electrophoresis , nucleic acid , microreactor , capillary action , chemistry , chromatography , enzyme , rna , electrophoresis , biochemistry , materials science , composite material , gene , catalysis
This report describes the coupling of capillary enzyme reactors to capillary electrophoresis, which is termed capillary enzymophoresis. In the present study, the capillary enzyme reactors were prepared by immobilizing RNA‐modifying enzymes, e.g. , RNase T 1 and RNase U 2 , on the inner walls of 50 μm fused‐silica capillaries. These microreactors served to selectively modify the solutes (or substrates) before entering the separation capillary. Capillary enzymophoresis using single or mixed enzyme reactors proved useful in identifying minute amounts of dinucleotides as well as the fingerprinting of tRNAs. The immobilized RNase T 1 and RNase U 2 displayed their usual enzymic activities toward RNA fragments and in addition exhibited different activity‐pH dependency than the soluble enzymes. This was attributed to microenvironmental effects arising from the charged nature of the capillary walls in the close proximity of the immobilized enzymes. The enzyme reactors were reusable for several RNA samples and showed chemical and thermal stability Presented as a part of a lecture (#313) at Pittsburgh Conference 94 (Pittcon '94), Chicago, Illinois, February 27‐March 4, 1994. .