Reloading and stability of polyacrylamide slab gels for automated DNA sequencing.
Current automated fluorescent instruments are all based on slab gels that are used once and then discarded. Practitioners of capillary gel electrophoresis often reuse their gels for multiple samples. As slab gels are made thinner to increase speed, the ability to reload new samples after each run will become more desirable. Techniques have previously been developed for reloading and stabilizing capillary gels. The application of these methods to slab gel electrophoresis is reported. Gels are shown to be reusable for at least four consecutive automated runs. The stability of various slab gel formulations and their ability to survive multiple loadings with sequencing samples are compared. Formamide-containing gels are shown to be superior to their urea counterparts. The potential that running buffer additives have for improving automated DNA sequencing is discussed. Residual template DNA in sequencing samples can produce gel instability, reduce resolution and decrease signal. These effects are examined.
University of Utah, Salt Lake City.
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Swerdlow
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Dew-Jager
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Gesteland
D001322:Autoanalysis / Q000379:methods*; D001483:Base Sequence; D002021:Buffers; D002197:Capillary Action; D004247:DNA / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D005559:Formamides; D010957:Plasmids; D017422:Sequence Analysis, DNA / Q000379:methods*; D013698:Templates, Genetic
D002021:Buffers; D004277:DNA, Single-Stranded; D005559:Formamides; C031066:formamide; D004247:DNA