A quick method for obtaining high-quality DNA barcodes without DNA extraction in microalgae

Abstract

The identification of microalgae based on their morphological features is always challenging due to their similar traits. DNA barcoding is an established technique for obtaining information about genomic sequences to identify different species. However, obtaining DNA from microalgae requires long cultivation times in order to produce enough cells and many steps are needed for the extraction of DNA, requiring significant time and money. In previous studies direct polymerase chain reaction (PCR) has been suggested as a feasible approach for some microalgae, but this protocol still requires pretreatment to deal with the specimens. In the current study, a direct PCR that avoids the DNA extraction step was tested in Chlorella, Scenedesmus, Dunaliella salina and diatoms living in freshwater and seawater. Our optimization involves the following three major aspects: (1) the use of different species, (2) protocol simplification, and (3) microalgae and DNA amount. We also detected DNA levels required for our direct PCR method. After the optimization, the improvement realized was our method’s simplicity, efficiency, and cost-effectiveness, since all the pretreatments were omitted. However, no alterations were observed in the results and equal quality PCR products were obtained compared with conventional PCR. Also, this technique has the same phylogenetic utility as the other PCR methods. To the best of our knowledge, this is the simplest and most direct way to perform PCR in both freshwater and marine microalgae. We also show that the optimized results can be used for routine multi-locus phylogenetic analysis and demonstrate high resolution of DNA barcoding gaps.