scholarly journals Development and Testing of Species-specific Quantitative PCR Assays for Environmental DNA Applications

10.3791/61825 ◽  
2020 ◽  
Author(s):  
Katy E. Klymus ◽  
Dannise V. Ruiz Ramos ◽  
Nathan L. Thompson ◽  
Catherine A. Richter
Keyword(s):  
Quantitative Pcr ◽  
Environmental Dna ◽  
Pcr Assays ◽  
Species Specific

scholarly journals Detection of four imperiled western North American freshwater mussel species from environmental DNA with multiplex qPCR assays

2020 ◽  
Author(s):  
Torrey W. Rodgers ◽  
Joseph C. Dysthe ◽  
Cynthia Tait ◽  
Thomas W. Franklin ◽  
Michael K. Schwartz ◽  
...  
Keyword(s):  
Freshwater Mussel ◽  
Environmental Dna ◽  
Vulnerable Species ◽  
Western North America ◽  
Mussel Species ◽  
Multiplex Assays ◽  
Specificity And Sensitivity ◽  
Pcr Assays ◽  
Species Specific ◽  
Management And Conservation

AbstractWe developed multiplexed, species-specific, quantitative PCR assays for the detection of four freshwater mussel species native to western North America, Gonidea angulata, Margaritifera falcata, Anodonta nuttalliana and Anodonta oregonensis, from environmental DNA (eDNA). These species have experienced dramatic declines over the last century and are currently threatened in many portions of their ranges. Therefore, improved tools for detecting and monitoring these species are needed. Species-specificity and sensitivity of assays were empirically tested in the lab, and multiplex assays were also validated with field collected eDNA samples. All assays were species-specific, sensitive, and effective for detection from eDNA samples collected from streams and rivers. These assays will aid in the detection, monitoring, management, and conservation of these vulnerable species.

scholarly journals Quantitative PCR assays for detection of five Arctic fish species: Lota lota, Cottus cognatus, Salvelinus alpinus, Salvelinus malma, and Thymallus arcticus from environmental DNA

2017 ◽  
Author(s):  
Torrey W. Rodgers ◽  
John. R. Olson ◽  
Stephen L. Klobucar ◽  
Karen. E. Mock
Keyword(s):  
Quantitative Pcr ◽  
Salvelinus Alpinus ◽  
Environmental Dna ◽  
North Slope ◽  
Cottus Cognatus ◽  
The North ◽  
North Slope Of Alaska ◽  
Arctic Fish ◽  
Thymallus Arcticus ◽  
Pcr Assays

AbstractThe North Slope of Alaska contains arctic fish populations that are important for subsistence of local human populations, and are under threat from natural resource extraction and climate change. We designed and evaluated four quantitative PCR assays for the detection of environmental DNA from five Alaskan fish species present on the North Slope of Alaska: burbot (Lota lota), arctic char (Salvelinus alpinus), Dolly Varden (Salvelinus malma), arctic grayling (Thymallus arcticus), and slimy sculpin (Cottus cognatus). All assays were designed and tested for species specificity and sensitivity, and all assays detected target species from filtered water samples collected from the field. These assays will enable efficient and economical detection and monitoring of these species in lakes and rivers. This in turn will provide managers with improved knowledge of current distributions and future range shifts associated with climate and development threats, enabling more timely management.

scholarly journals Development of a Quantitative PCR Assay for Four Salmon Species Inhabiting the Yangyangnamdae River Using Environmental DNA

Biology ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 899
Author(s):  
Muhammad Hilman Fu'adil Amin ◽  
Ji-Hyun Lee ◽  
Ah Ran Kim ◽  
Ju-Kyoung Kim ◽  
Chung-Il Lee ◽  
...  
Keyword(s):  
Quantitative Pcr ◽  
Native Species ◽  
Limit Of Detection ◽  
Environmental Dna ◽  
Qpcr Assay ◽  
Spawning Season ◽  
Oncorhynchus Keta ◽  
Korean Waters ◽  
Quantitative Analyses ◽  
Species Specific

A species-specific quantitative PCR (qPCR) assay using environmental DNA (eDNA) is a promising tool for both qualitative and quantitative analyses of target species directly from water samples. Despite its reliability, an eDNA-based qPCR assay pipeline has not yet developed to monitor salmon species inhabiting Korean waters, which have been rapidly decreasing. We designed species-specific primers for four Oncorhynchus species inhabiting the eastern coastal waters along the Korean Peninsula. These include primers for two native species (Oncorhynchus keta and O. masou) and two that were introduced (O. mykiss and O. kisutch). The limit of detection and limit of quantification for the four qPCR assays ranged from 4.11 to 10.38 copies and from 30 to 81 copies, respectively, indicating a high sensitivity and specificity across all four species. Following optimization, the qPCR assays were used for the quantitative analyses of the four Oncorhynchus species in the Yangyangnamdae River during the spawning and non-spawning seasons in the year 2019–2020, one of the main rivers where salmon migrate during the spawning season in Korea. The raw copy numbers in all of the examined samples were normalized by PCR inhibition rates to standardize and compare with other studies. Among the four Oncorhynchus species examined, the eDNA concentration of O. keta increased significantly (63.60-fold, p < 0.0001) during the spawning season (November) compared with that in the non-spawning season (March), suggesting that O. keta is the main salmon species migrating through the Yangyangnamdae River. In contrast, we did not detect any differences in eDNA concentration for the other three Oncorhynchus species between the spawning and non-spawning seasons, indicating that their presence does not alter during the year. Their eDNA concentration is also relatively low compared to O. keta, which suggests that small numbers of these three species are present in the river. Overall, these newly developed qPCR assays represent useful monitoring tools for the management of four salmon species in Korean waters.

scholarly journals A quantitative PCR based environmental DNA assay for detecting Atlantic salmon (Salmo salar L.)

2017 ◽  
Author(s):  
Siobhán Atkinson ◽  
Jeannette E.L. Carlsson ◽  
Bernard Ball ◽  
Damian Egan ◽  
Mary Kelly-Quinn ◽  
...  
Keyword(s):  
Atlantic Salmon ◽  
Quantitative Pcr ◽  
Salmo Salar ◽  
Environmental Dna ◽  
Coi Gene ◽  
List Type ◽  
The Pacific ◽  
Invasive Method ◽  
Species Specific ◽  
A Minor

AbstractThe Atlantic salmon (Salmo salar L.) has worldwide ecological, cultural and economic importance. The species has undergone extensive decline across its native range, yet concerns have been raised about its invasive potential in the Pacific. Knowledge on the distribution of this species is vital for addressing conservation goals.This study presents an eDNA assay to detect S. salar in water samples, using quantitative PCR (qPCR) technology. Species-specific primers and a minor groove binding (MGB) probe were designed for the assay, based on the mitochondrial cytochrome oxidase I (COI) gene.The results of this study indicate that eDNA is a highly sensitive tool for detecting S. salar in situ, and could potentially provide an alternative, non-invasive method for determining the distribution of this species.

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