Assessment of EvaGreen-based quantitative real-time PCR assay for enumeration of the microalgae Heterosigma and Chattonella (Raphidophyceae) SCIE SCOPUS

Cited 14 time in WEB OF SCIENCE Cited 14 time in Scopus
Title
Assessment of EvaGreen-based quantitative real-time PCR assay for enumeration of the microalgae Heterosigma and Chattonella (Raphidophyceae)
Author(s)
Park, Bum Soo; Baek, Seung Ho; Ki, Jang-Seu; Cattolico, Rose Ann; Han, Myung-Soo
KIOST Author(s)
Baek, Seung Ho(백승호)
Publication Year
2012-12
Abstract
Heterosigma akashiwo and Chattonella species (Raphidophyceae) are difficult to detect and quantify in environmental samples because of their pleomorphic and fragile cell nature. In this study, we developed a quantitative real-time polymerase chain reaction (qRT-PCR) assay for the enumeration of these algal taxa using a new DNA-binding dye, EvaGreen. Species-specific qRT PCR primers to H. akashiwo, Chattonella antiqua, Chattonella marina, Chattonella ovata, and Chattonella subsalsa were designed to target the ITS2 rRNA gene intergenic region. Primer specificities were tested via BLAST searches. In addition, specificity was verified using empirical tests, including competitive PCR. The qRT PCR assay analyzing C (t) value and the log of cell number showed a significant linear relationship (r (2) a parts per thousand yenaEuro parts per thousand 0.997). When light microscopy was used to monitor the population dynamics of targeted Raphidophyceae from Lake Shihwa, H. akashiwo was detected in ten samples and no Chattonella spp. were detected (70 samples collected from May, 2007 to January, 2008). In contrast, when the qRT-PCR assay was used, H. akashiwo was detected in 41 samples. C. antiqua, C. marina, and C. ovata were detected in eight samples. Most of the samples analyzed using qRT-PCR assays showed higher algal numbers than did those assayed via microscopy, suggesting that the enumeration of Raphidophyceae via classic microscopic methods most likely underestimates true algal concentration.
ISSN
0921-8971
URI
https://sciwatch.kiost.ac.kr/handle/2020.kiost/3420
DOI
10.1007/s10811-012-9816-2
Bibliographic Citation
JOURNAL OF APPLIED PHYCOLOGY, v.24, no.6, pp.1555 - 1567, 2012
Publisher
SPRINGER
Subject
POLYMERASE-CHAIN-REACTION; RIBOSOMAL-RNA GENE; SYBR-GREEN-I; PARASITE PERKINSUS-MARINUS; MELTING CURVE ANALYSIS; RED TIDE ALGAE; PFIESTERIA-PISCICIDA; ENVIRONMENTAL-SAMPLES; RAPID DETECTION; DNA FRAGMENTS
Keywords
Raphidophyceae; Quantitative real-time PCR; EvaGreen; Field application
Type
Article
Language
English
Document Type
Article
Publisher
SPRINGER
Related Researcher
Research Interests

Marine Ecology,Marine Environment,HABs,해양생태,해양환경,적조

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