Automation of the Maxam-Gilbert chemical sequencing reactions.
A practical automated method of Maxam-Gilbert chemical sequencing reactions that uses solid-phase chromatography methods to purify DNA following chemical modification and cleavage is described in this report. The automation has primarily been made possible by using specially designed BioPak mini-columns, compatible with the Biomek 1000 automated workstation, which can be utilized in a manner similar to that of standard pipet tips. This automated chromatographic sequencing method produces rapid and reliable data as verified by sequencing a known human factor IX exon VIII gene fragment. The procedure presented in this report is a prototype for a single-fragment reaction and can easily be expanded to perform reactions on as many as 8 fragments at a time. The automation eliminates the tedious and time-consuming steps in the original method and increases the rate of sequence acquisition. This technology makes the Maxam-Gilbert chemical sequencing protocol more accessible, especially in large-scale, automated sequencing projects.
Boland
E J
EJ
University of Texas Health Science Center at San Antonio 78284-7762.
Pillai
A
A
Odom
M W
MW
Jagadeeswaran
P
P
D001322:Autoanalysis / Q000379:methods*; D001345:Autoradiography; D001483:Base Sequence; D002845:Chromatography / Q000379:methods; D004247:DNA / Q000737:chemistry / Q000302:isolation & purification; D005091:Exons; D005164:Factor IX / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D017422:Sequence Analysis, DNA
D005164:Factor IX; D004247:DNA