DNA extraction, preservation, and amplification.
The effectiveness of DNA barcoding as a routine practice in biodiversity research is strongly dependent on the quality of the source material, DNA extraction method, and selection of adequate primers in combination with optimized polymerase chain reaction (PCR) conditions. For the isolation of nucleic acids, silica-gel membrane methods are to be favored because they are easy to handle, applicable for high sample throughput, relatively inexpensive, and provide high DNA quality, quantity, and purity which are pre-requisites for successful PCR amplification and long-term storage of nucleic acids in biorepositories, such as DNA banks. In this section, standard protocols and workflow schemes for sample preparation, DNA isolation, DNA storage, PCR amplification, PCR product quality control, and PCR product cleanup are proposed and described in detail. A PCR troubleshooting and primer design section may help to solve problems that hinder successful amplification of the desired barcoding gene region.
German Centre for Marine Biodiversity Research (DZMB), Senckenberg Research Institute, Wilhelmshaven, Germany. tknebelsberger@senckenberg.de
Thomas
T
Knebelsberger
Isabella
I
Stöger
D004247:DNA / Q000235:genetics* / Q000302:isolation & purification*; D058893:DNA Barcoding, Taxonomic; D016133:Polymerase Chain Reaction
D004247:DNA