Analysis and purification of synthetic DNA fragments with NuSieve agarose mini-gels.
We outline in this note a new method for analyzing the crude product from long (greater than 40 bp) DNA synthesis reactions. This procedure is quick, and provides direct visualization of the end product. As described here, this method is not quantitative. However, densitometry of the stained gel could be used to quantitate the amount of DNA in the full-length peak. Without quantitation, our experience indicates that if a full-length fragment in the size range of 60 to 106 bases cannot be visualized with up to 1 microgram of crude material, re-synthesis is indicated. The 1 microgram value will no doubt increase with longer fragments. This system has also been used to purify ligated fragments for cloning. We have not attempted to purify full-length single-stranded oligomers from the crude reaction mix using the agarose mini-gel system, because of the lack of resolution at the n-1 level, and also because of the small loading capacity of the gel.
Zupancic
T J
TJ
Biotechnology Dept., Battelle Columbus Labs, Columbus, OH 43201.
Hilt
D A
DA
Zarley
C D
CD
Kimball
P C
PC
D004247:DNA / Q000032:analysis* / Q000138:chemical synthesis / Q000302:isolation & purification; D004586:Electrophoresis / Q000379:methods*; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D012685:Sepharose / Q000031:analogs & derivatives
C064154:NuSieve agarose; D004247:DNA; D012685:Sepharose