Abstract
A simple miniaturized gel system suitable for DNA sequencing is described. Small ultrathin polyacrylamide gels are cast, eight or more at a time, using standard microscope slides. Gels, ready to use, can be stored for approximately 2 weeks. Gels are run horizontally in a standard mini-agarose gel apparatus. Typical run times are 6-8 min. A novel sample loading system permits volumes of standard sequencing reactions as small as 0.1 microl to be analyzed. Sequencing ladders were visualized using 35S-labeled DNA by autoradiography and by colorimetric detection. Band resolution compares favorably with that of large gels. The methods introduced here serve as a step toward the miniaturization of DNA sequencing and are amenable to automated sample loading and detection.
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Selected References
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- Ansorge W., Barker R. System for DNA sequencing with resolution of up to 600 base pairs. J Biochem Biophys Methods. 1984 Mar;9(1):33–47. doi: 10.1016/0165-022x(84)90064-2. [DOI] [PubMed] [Google Scholar]
- Brumley R. L., Jr, Smith L. M. Rapid DNA sequencing by horizontal ultrathin gel electrophoresis. Nucleic Acids Res. 1991 Aug 11;19(15):4121–4126. doi: 10.1093/nar/19.15.4121. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Drossman H., Luckey J. A., Kostichka A. J., D'Cunha J., Smith L. M. High-speed separations of DNA sequencing reactions by capillary electrophoresis. Anal Chem. 1990 May 1;62(9):900–903. doi: 10.1021/ac00208a003. [DOI] [PubMed] [Google Scholar]
- Garoff H., Ansorge W. Improvements of DNA sequencing gels. Anal Biochem. 1981 Aug;115(2):450–457. doi: 10.1016/0003-2697(81)90031-2. [DOI] [PubMed] [Google Scholar]
- Richterich P., Church G. M. DNA sequencing with direct transfer electrophoresis and nonradioactive detection. Methods Enzymol. 1993;218:187–222. doi: 10.1016/0076-6879(93)18016-6. [DOI] [PubMed] [Google Scholar]
- Swerdlow H., Gesteland R. Capillary gel electrophoresis for rapid, high resolution DNA sequencing. Nucleic Acids Res. 1990 Mar 25;18(6):1415–1419. doi: 10.1093/nar/18.6.1415. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Woolley A. T., Mathies R. A. Ultra-high-speed DNA sequencing using capillary electrophoresis chips. Anal Chem. 1995 Oct 15;67(20):3676–3680. doi: 10.1021/ac00116a010. [DOI] [PubMed] [Google Scholar]
