A DNA Isolation Protocol Suitable for RAPD Analysis from Fresh or Herbarium-Stored Leaves of a Historic Quercus virginiana L.

摘要

This study described a fast, high-quality yielding protocol for the extraction of genomic DNA from either fresh or herbarium-stored plant tissue of an historic Quercus virginiana called the ?Emancipation Oak?. The protocol was based on the combination of two existing methods-CTAB (hexadecyltrimethylammonium bromide) and DNAzol. The combined DNAzol?-CTAB (C-DC) procedure avoids the traditional phenol: chloroform extraction which was needed to remove polysaccharides and proteins that were normally found in DNA isolated from plants. The C-DC protocol was compared to two standard methodologies for the isolation of intact DNA from either fresh or herbarium-stored plant tissue. The C-DC methodology yields final DNA (from either fresh or herbarium-stored samples) that can be used for RAPD analysis without the need to treat with RNAase. Differences in DNA yields among the three protocols with either fresh or herbarium-stored tissue were analyzed via an ANOVA. The isolated C-DC DNA was free of polysaccharides, polyphenols, and other major contaminants as judged by A260/280 ratio, 0.8% agarose gel electrophoresis and RAPD suitability. This new C-DC protocol will be of great help to study the genetic diversity of the ?Emancipation Oak?,as well as other species of Quercus .