Nucleic Acids Research, 1985, Vol. 13, No. 4 1173-1184
© 1985
Articles |
Solid-phase methods for sequencing of nucleic acids I. Simultaneous sequencing of different oligodeoxyribonucleotides using a new, mechanically stable anion-exchange paper+
Academy of Sciences of GDR, Central Institute of Molecular Biology, Department of Cell Differentiation Robert-Rössle-Strasse 10, 1115 Berlin-Buch, GDR
To whom all correspondence should be addressed
Received December 4, 1984. Revised January 24, 1985. Accepted January 24, 1985.
A solid-phase method for simultaneous sequencing of large numbers of oligodeoxyribonucleotidea has been developed using a new, mechanically stable anion-exchange paper. The excellent mechanical properties of the polymer allow the processing of several paper segments in one reaction vessel or to carry out all necessary operations on a larger area of the paper. In addition, DNA material can be chemically eluted from the new carrier during the piperidine reaction, thus avoiding salt elution of DNA and subsequent ethanol precipitation steps - a prerequisite for sequencing oligonucleotides. The approach involves 7 operations including: i) immobilization; ii) washing; iii) modification; iv) washing; v) sorting of the papers; vi) piperidine reaction and chemical elution and vii) lyophilization. All steps can be carried out in 4 to 5 hours independently of the number of oligonucleotides to be sequenced. It is also possible to sequence small oligonucleotides with 3 to 4 base pairs. The method can be fully automated.
+A part of this paper hss been presented at the VI. Symposium on the Chemistry of Nucleic Acid Components in Prague, September 2nd-8th, 1984.