Premium
Separation of 4‐color DNA sequencing extension products in noncovalently coated capillaries using low viscosity polymer solutions
Author(s) -
Mandabhushi Ramakrishna S.
Publication year - 1998
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.1150190215
Subject(s) - capillary action , capillary electrophoresis , viscosity , coating , polymer , chromatography , chemistry , electrophoresis , capillary length , covalent bond , resist , analytical chemistry (journal) , materials science , chemical engineering , nanotechnology , layer (electronics) , composite material , organic chemistry , engineering
A low viscosity ( ca. 75cp) solution using polydimethylacrylamide (PDMA) was developed for separating DNA sequencing extension products by capillary electrophoresis (CE). This medium gave a length‐of‐read (LOR) value of approximately 600 bases in about 2 h using four‐color sequencing in 50 μm capillary at 42°C under a field of 160 V/cm. This medium also works in bare capillaries by noncovalently coating the surface to suppress both electroosmotic flow (EOF) and DNA‐capillary wall interactions, and eliminates the need for complicated covalent coatings. At least 100 successive sequencing runs were performed in the same capillary by simply pumping fresh medium after every run, without requiring any reconditioning of the capillary surface between runs. The thermal stability of the noncovalent coating can be improved by adding small amounts of high molecular weight PDMA to the separation medium. The advantages of low viscosity separation media and uncoated capillaries are of paramount importance to develop high‐throughput instruments for DNA sequencing.