분석조건별 담수어류의 환경 DNA 메타바코딩 효율 비교: 필터, 추출 키트, 프라이머 조합 및 PCR 방법 (Efficiency Comparison of Environmental DNA Metabarcoding of Freshwater Fishes according to Filters, Extraction Kits, Primer Sets and PCR Methods.)

서지정보

· 발행기관 : 한국수생태학회
· 수록지 정보 : 생태와 환경 / 54권 / 3호 / 199 ~ 208페이지
· 저자명 : 김근식, 김근용, 윤주덕

초록

Environmental DNA (eDNA) metabarcoding is effective method with high detection sensitivity for evaluating fish biodiversity and detecting endangered fish from natural water samples. We compared the richness of operational taxonomic units (OTUs) and composition of freshwater fishes according to filters (cellulose nitrate filter vs. glass fiber filter), extraction kits (DNeasy2® Blood & Tissue Kit vs. DNeasy2® PowerWater Kit), primer sets (12S rDNA vs. 16S rDNA), and PCR methods (conventional PCR vs. touchdown PCR) to determine the optimal conditions for metabarcoding analysis of Korean freshwater fish. The glass fiber filter and DNeasy2® Blood & Tissue Kit combination showed the highest number of freshwater fish OTUs in both 12S and 16S rDNA. Among the four types, the primer sets only showed statistically significant difference in the average number of OTUs in class Actinopterygii (non-parametric Wilcoxon signed ranks test, p = 0.005). However, there was no difference in the average number of OTUs in freshwater fish. The species composition also showed significant difference according to primer sets (PERMANOVA, Pseudo-F = 6.9489, p = 0.006), but no differences were observed in the other three types. The non-metric multidimensional scaling (NMDS) results revealed that species composition clustered together according to primer sets based on similarity of 65%; 16S rDNA primer set was mainly attributed to endangered species such as Microphysogobio koreensis and Pseudogobio brevicorpus. In contrast, the 12S rDNA primer set was mainly attributed to common species such as Zacco platypus and Coreoperca herzi. This study provides essential information on species diversity analysis using metabarcoding for environmental water samples obtained from rivers in Korea.

영어초록

Environmental DNA (eDNA) metabarcoding is effective method with high detection sensitivity for evaluating fish biodiversity and detecting endangered fish from natural water samples. We compared the richness of operational taxonomic units (OTUs) and composition of freshwater fishes according to filters (cellulose nitrate filter vs. glass fiber filter), extraction kits (DNeasy2® Blood & Tissue Kit vs. DNeasy2® PowerWater Kit), primer sets (12S rDNA vs. 16S rDNA), and PCR methods (conventional PCR vs. touchdown PCR) to determine the optimal conditions for metabarcoding analysis of Korean freshwater fish. The glass fiber filter and DNeasy2® Blood & Tissue Kit combination showed the highest number of freshwater fish OTUs in both 12S and 16S rDNA. Among the four types, the primer sets only showed statistically significant difference in the average number of OTUs in class Actinopterygii (non-parametric Wilcoxon signed ranks test, p = 0.005). However, there was no difference in the average number of OTUs in freshwater fish. The species composition also showed significant difference according to primer sets (PERMANOVA, Pseudo-F = 6.9489, p = 0.006), but no differences were observed in the other three types. The non-metric multidimensional scaling (NMDS) results revealed that species composition clustered together according to primer sets based on similarity of 65%; 16S rDNA primer set was mainly attributed to endangered species such as Microphysogobio koreensis and Pseudogobio brevicorpus. In contrast, the 12S rDNA primer set was mainly attributed to common species such as Zacco platypus and Coreoperca herzi. This study provides essential information on species diversity analysis using metabarcoding for environmental water samples obtained from rivers in Korea.

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