Abstract

Genomic DNA is beneficial for nematode study which provides sufficient information for various purposes. Nowadays, DNA-based techniques are used as a supplementary method to identify species of this genus and phylogenetic studies more accurately and efficiently. However, DNA extraction of plant-parasitic is challenging. Additionally, some methods such as chloroform are hazardous. Helicotylenchus is one of the critical plant-parasites nematodes that cause damage to agricultural products. This study was conducted from March to April 2019 at the University of Limpopo, South Africa to evaluate two DNA extraction methods' efficiency. Helicotylenchus sp. was collected from the grass in a garden and identified using the conventional method. Besides in a method, after transferring live nematodes (5, 10, and 20 alive individuals) to the Eppendorf sterile tubes containing 16 μL twice sterile distilled water, the nematodes were crushed by a sterile needle and then the tubes were immersed in 100°C boiling water for 10 minutes. In this method, no proteinase K was used. After DNA extraction, the samples were tested using D2-D3 primers of the 28S rDNA genomic region. Furthermore, the PCR product showed that DNA extraction from Helicotylenchus sp. using 10 and 20 individuals yielded the DNA for PCR processing toward species identification. Additionally, the other method using proteinase K also was used for DNA extraction which the result showed sufficient quality of the extracted DNA. DNA can be extracted in both methods for Helicotylenchus, however, when the proteinase K is not available, DNA extraction using boiling water with enough individuals can be used.