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Volume 13
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10.3390/genes13020175
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Article

Selection of References for microRNA Quantification in Japanese Flounder (Paralichthys olivaceus) Normal Tissues and Edwardsiella tarda-Infected Livers

by
Saisai Liu
1,
Haofei Song
1,
Zeyu Liu
1,
Wei Lu
1,
Quanqi Zhang
1,2,3 and
Jie Cheng
1,2,3,*
1
Key Laboratory of Marine Genetics and Breeding, Ocean University of China, Ministry of Education, 5 Yushan Road, Qingdao 266003, China
2
Laboratory for Marine Fisheries Science and Food Production Processes, Pilot National Laboratory for Marine Science and Technology (Qingdao), 1 Wenhai Road, Qingdao 266237, China
3
Laboratory of Tropical Marine Germplasm Resources and Breeding Engineering, Sanya Oceanographic Institution, Ocean University of China, Sanya 572024, China
*
Author to whom correspondence should be addressed.
Genes 2022, 13(2), 175; https://doi.org/10.3390/genes13020175
Submission received: 27 December 2021 / Revised: 9 January 2022 / Accepted: 17 January 2022 / Published: 19 January 2022
(This article belongs to the Special Issue Genomics in Aquaculture and Fisheries)
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Abstract

MicroRNA (miRNA) plays essential roles in post-transcriptional regulation of protein coding genes, and the quantitative real-time polymerase chain reaction (qRT-PCR) is the powerful and broadly employed tool to conduct studies of miRNA expression. Identifying appropriate references to normalize quantitative data is a prerequisite to ensure the qRT-PCR accuracy. Until now, there has been no report about miRNA reference for qRT-PCR in Japanese flounder (Paralichthys olivaceus), one important marine cultured fish along the coast of Northern Asia. In this study, combined with miRNA-Seq analysis and literature search, 10 candidates (miR-34a-5p, miR-205-5p, miR-101a-3p, miR-22-3p, miR-23a-3p, miR-210-5p, miR-30c-5p, U6, 5S rRNA, and 18S rRNA) were chosen as potential references to test their expression stability among P. olivaceus tissues, and in livers of P. olivaceus infected with Edwardsiella tarda at different time points. The expression stability of these candidates was analyzed by qRT-PCR and evaluated with Delta CT, BestKeeper, geNorm, as well as NormFinder methods, and RefFinder was employed to estimate the comprehensive ranking according to the four methods. As the result, miR-22-3p and miR-23a-3p were proved to be the suitable combination as reference miRNAs for both P. olivaceus normal tissues and livers infected with E. tarda, and they were successfully applied to normalize miR-7a and miR-221-5p expression in P. olivaceus livers in response to E. tarda infection. All these results provide valuable information for P. olivaceus miRNA quantitative expression analysis in the future.
Keywords: microRNA; reference gene; qRT-PCR; Paralichthys olivaceus; Edwardsiella tarda microRNA; reference gene; qRT-PCR; Paralichthys olivaceus; Edwardsiella tarda

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MDPI and ACS Style

Liu, S.; Song, H.; Liu, Z.; Lu, W.; Zhang, Q.; Cheng, J. Selection of References for microRNA Quantification in Japanese Flounder (Paralichthys olivaceus) Normal Tissues and Edwardsiella tarda-Infected Livers. Genes 2022, 13, 175. https://doi.org/10.3390/genes13020175

AMA Style

Liu S, Song H, Liu Z, Lu W, Zhang Q, Cheng J. Selection of References for microRNA Quantification in Japanese Flounder (Paralichthys olivaceus) Normal Tissues and Edwardsiella tarda-Infected Livers. Genes. 2022; 13(2):175. https://doi.org/10.3390/genes13020175

Chicago/Turabian Style

Liu, Saisai, Haofei Song, Zeyu Liu, Wei Lu, Quanqi Zhang, and Jie Cheng. 2022. "Selection of References for microRNA Quantification in Japanese Flounder (Paralichthys olivaceus) Normal Tissues and Edwardsiella tarda-Infected Livers" Genes 13, no. 2: 175. https://doi.org/10.3390/genes13020175

APA Style

Liu, S., Song, H., Liu, Z., Lu, W., Zhang, Q., & Cheng, J. (2022). Selection of References for microRNA Quantification in Japanese Flounder (Paralichthys olivaceus) Normal Tissues and Edwardsiella tarda-Infected Livers. Genes, 13(2), 175. https://doi.org/10.3390/genes13020175

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