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Int J Mol Sci. 2017 Jan 4;18(1). pii: E84. doi: 10.3390/ijms18010084.

Detection of Ribosomal DNA Sequence Polymorphisms in the Protist Plasmodiophora brassicae for the Identification of Geographical Isolates.

Author information

1
Department of Horticulture, Sunchon National University, Suncheon 540-950, Korea. rawnak.2010@gmail.com.
2
Department of Horticulture, Sunchon National University, Suncheon 540-950, Korea. gpb21bau@gmail.com.
3
Department of Horticulture, Sunchon National University, Suncheon 540-950, Korea. ykw7685@naver.com.
4
Center for Eco-friendly New Materials, Korea Research Institute of Chemical Technology, Daejeon 34114, Korea. kjchoi@krict.re.kr.
5
Department of Horticulture, Sunchon National University, Suncheon 540-950, Korea. jipark@sunchon.ac.kr.
6
Department of Horticulture, Sunchon National University, Suncheon 540-950, Korea. nis@sunchon.ac.kr.

Abstract

Clubroot is a soil-borne disease caused by the protist Plasmodiophora brassicae (P. brassicae). It is one of the most economically important diseases of Brassica rapa and other cruciferous crops as it can cause remarkable yield reductions. Understanding P. brassicae genetics, and developing efficient molecular markers, is essential for effective detection of harmful races of this pathogen. Samples from 11 Korean field populations of P. brassicae (geographic isolates), collected from nine different locations in South Korea, were used in this study. Genomic DNA was extracted from the clubroot-infected samples to sequence the ribosomal DNA. Primers and probes for P. brassicae were designed using a ribosomal DNA gene sequence from a Japanese strain available in GenBank (accession number AB526843; isolate NGY). The nuclear ribosomal DNA (rDNA) sequence of P. brassicae, comprising 6932 base pairs (bp), was cloned and sequenced and found to include the small subunits (SSUs) and a large subunit (LSU), internal transcribed spacers (ITS1 and ITS2), and a 5.8s. Sequence variation was observed in both the SSU and LSU. Four markers showed useful differences in high-resolution melting analysis to identify nucleotide polymorphisms including single- nucleotide polymorphisms (SNPs), oligonucleotide polymorphisms, and insertions/deletions (InDels). A combination of three markers was able to distinguish the geographical isolates into two groups.

KEYWORDS:

Plasmodiophora brassicae; geographical isolates; intraspecific polymorphism; ribosomal DNA; sequence variation; single-nucleotide polymorphism

PMID:
28054984
PMCID:
PMC5297718
DOI:
10.3390/ijms18010084
[Indexed for MEDLINE]
Free PMC Article

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