2Vavilov Institute of General Genetics, Russian Academy of Sciences, 119333 Moscow, Russia; fax: +7 (499) 132-8962
3Central Research Institute of Epidemiology, Federal Service on Customers Rights Protection and Human Well-being Surveillance, 111123 Moscow, Russia; fax: +7 (495) 304-2209
* To whom correspondence should be addressed.
Received July 6, 2015; Revision received July 24, 2015
Here we report a rapid and cost-effective method for the extraction of total DNA from herbarium specimens up to 50-90-year-old. The method takes about 2 h, uses AMPure XP magnetic beads diluted by PEG-8000-containing buffer, and does not require use of traditional volatile components like chloroform, phenol, and liquid nitrogen. It yields up to 4 µg of total nucleic acid with high purity from about 30 mg of dry material. The quality of the extracted DNA was tested by PCR amplification of 5S rRNA and rbcL genes (nuclear and chloroplast DNA markers) and compared against the traditional chloroform/isoamyl alcohol method. Our results demonstrate that the use of the magnetic beads is crucial for extraction of DNA suitable for subsequent PCR from herbarium samples due to the decreasing inhibitor concentrations, reducing short fragments of degraded DNA, and increasing median DNA fragment sizes.
KEY WORDS: DNA extraction, herbarium, PCR, 5S rRNA, rbcL, genomic markers, sequence