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Detection and Identification Root Knot Nematode (Meloidogyne spp.) in Sugar Beet Using DNA Barcoding Method
Nematode identification based on morphological characters requires high accuracy and needs to be supported by molecular identification results. One method of molecular identification is DNA barcodes. DNA barcode is a molecular identification using the cythochrome oxidase sub unit 1 (CO1) gene. This study aims to identify NPA associated with beetroot based on PCR and Meloidogyne CO1 nucleotide gene sequencing. Beetroot samples were obtained from agricultural land located in Cipanas District, Cianjur Regency, West Java. Sampling was done by purposive sampling method. Morphological identification by observing the perineal pattern of female NPA. Molecular identification includes extraction, amplification, DNA visualization, nucleotide and phylogenetic analysis. Morphological perineal pattern observations obtained M. incognita and M. arenaria species. DNA amplification was carried out using CO1SIF (5’-GCCTGCATTTGGTTAG-‘3) and CO1SIR (5’-TCAAACCAGTCCT-‘3), CO1SAF (5’-GGGTACTGGATGAACATTA-‘3) and CO1SAR (5’-ACTTCAGGATGACCAAA-‘3). DNA amplification succeeded in getting a DNA band measuring ± 360 for M. arenaria and ± 326 for M. incognita. The sequencing results showed that M. incognita isolates from Indonesia were closely related to M. incognita isolates from China, US, UK, and Vietnam with a 100% homology level. The results of the phylogeny analysis showed that M. incognita Indonesia was still in the same group as M. incognita from China, US, UK, and Vietnam. M. arenaria isolates were closely related to M. arenaria isolates from Argentina and US with a homology level of 100%. The results of phylogenetic analysis showed that M. arenaria from Indonesia was still in the same group as M. arenaria from Argentina and US.
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