ORIGINAL ARTICLE
Optimization of DNA extraction methods for genomic analysis of rice root-knot nematode (Meloidogyne graminicola) using PCR (polymerase chain reaction) and sanger sequencing
1
Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
A - Research concept and design; B - Collection and/or assembly of data; C - Data analysis and interpretation; D - Writing the article; E - Critical revision of the article; F - Final approval of article
Submission date: 2022-08-26
Acceptance date: 2022-10-17
Online publication date: 2023-01-09
Corresponding author
Siwi Indarti
Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
Plant Protection, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
Journal of Plant Protection Research 2023;63(1):50–58
HIGHLIGHTS
- One nematode can be used for DNA extraction by freezing treatment before extraction.
- The commercial kit method is recommended for quick results of nematode DNA extraction.
- CTAB and SDS methods are recommended if the time is available and costs are limited.
KEYWORDS
TOPICS
ABSTRACT
The root-knot nematode Meloidogyne graminicola is an economically important pest in rice
production. The identification of a nematode species is an important basis in nematode
management to reduce yield losses by extracting nematode DNA as an early step in molecular
identification. This study aimed to investigate the optimal extraction method and
number of M. graminicola for nematode genomic analysis based on PCR (polymerase chain
reaction) and Sanger sequencing. The DNA extraction methods used in this study were the
CTAB, SDS, and commercial kit (GeneAidTM Tissue/Blood DNA Mini Kit). The results
revealed that the three DNA extraction methods could be used to analyze the nematode genomics
based on PCR and Sanger sequencing using one nematode, both in a second-stage
juvenile and a female, equipped with the process of nematode destruction by freezing. This
finding was shown by the amplification of all DNA templates with Mg-F3 and Mg-R2 primers
through PCR with a size of 370 bp, while Sanger sequencing obtained 372 bp.
ACKNOWLEDGEMENTS
The authors would like to thank Gadjah Mada University
for research funded by RTA (Rekognisi Tugas
Akhir-Student Final Project Recognition) with Contract
No. 3550/UN1.P.III/Dit-Lit/PT.01.05/2022.
RESPONSIBLE EDITOR
Renata Dobosz
CONFLICT OF INTEREST
The authors have declared that no conflict of interests exist.
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