A rapid genomic DNA extraction method and its combination with helicase dependent amplification for the detection of genetically modified maize

Abstract

It is compulsory for many crop plants to be tested for genetic modifications when entering the European Union. In this regard, a DNA extraction is performed and later a qualitative or quantitative PCR to detect and quantify any artificial inserts into the genome. Unfortunately, PCR needs specialized equipment and skilled personnel. This disadvantage has been solved by using isothermal DNA amplification methods, which are as sensitive as the PCR itself and can be performed at constant temperatures and therefore PCR thermocyclers are not needed. Nonetheless, those methods require high quality DNA and the isolation of it is still considered to be an elaborate process. Conventional DNA extraction methods are highly time consuming and tedious. In addition, they cannot be performed on-site as a centrifuge is always required. With this work, the development of a rapid method for DNA extraction from maize was carried out to overcome these problems. The method is based on the employ of an aqueous buffer system in combination with a proteinase K digestion and followed by a later filtration over a polypropylene membrane. Detection was carried out by helicase dependent amplification, as an alternative to PCR for the detection of transgenic maize. Data obtained are similar to those achieved with the more complex standard CTAB extraction method or the Promega Wizard DNA purification kit. The proposed DNA extraction method can be performed on-site, is inexpensive, simple and time saving.