[논문]RAG-1 and IgM Genes, Markers for Early Development of the Immune System in Olive Flounder, Paralichthys olivaceus

Lee, Jang-Wook; Yang, Hyun; Noh, Jae Koo; Kim, Hyun Chul; Park, Choul-Ji; Park, Jong-Won; Hwang, In Joon; Kim, Sung Yeon; Lee, Jeong-Ho

doi:10.12717/dr.2014.18.2.099

RAG-1 and IgM Genes, Markers for Early Development of the Immune System in Olive Flounder, Paralichthys olivaceus 원문보기

Development & reproduction = 발생과 생식, v.18 no.2, 2014년, pp.99 - 106

Lee, Jang-Wook (Genetics and Breeding Research Center, NFRDI) , Yang, Hyun (Genetics and Breeding Research Center, NFRDI) , Noh, Jae Koo (Genetics and Breeding Research Center, NFRDI) , Kim, Hyun Chul (Genetics and Breeding Research Center, NFRDI) , Park, Choul-Ji (Genetics and Breeding Research Center, NFRDI) , Park, Jong-Won (Genetics and Breeding Research Center, NFRDI) , Hwang, In Joon (Genetics and Breeding Research Center, NFRDI) , Kim, Sung Yeon (Genetics and Breeding Research Center, NFRDI) , Lee, Jeong-Ho (Genetics and Breeding Research Center, NFRDI)

Abstract ▼ AI-Helper

Fish larvae are immediately exposed to microbes from hatching to maturation of their lymphoid organs, therefore effective innate mechanisms is very important for survival. However, the knowledge of the development of immune system in fish is limited and in demand now. In vertebrates, recombination-activating gene 1 (RAG-1) and immunoglobulin M (IgM) have been considered as very useful markers of the physiological maturity of the immune system. In this study, the expression of the both genes was assessed throughout the early developmental stages of olive flounder larvae (5-55 dph) and used as markers to follow the development of immune system. RAG-1 and IgM mRNA expression was detectable at 5 dph and remained so until 55 dph. These patterns of expression may suggest that the olive flounder start to develop its function around 5 dph. Tissue distribution was found that both genes mRNAs are only expressed in the immune-related organ such as spleen, kidney and gill. The early detection of IgM mRNA led to the investigation of its presence in oocytes. Both RAG-1 and IgM mRNA transcripts were detected in unfertilized oocytes, suggesting that they are maternally transferred. The biological significance of such a phenomenon remains to be investigated.

주제어

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문제 정의

Because of the importance of immune response, many studies have been conducted in fish during the early larvae stages. This study is the first to report the mRNA expression of the important immune genes RAG-1 and IgM in the olive flounder. In addition their expression was followed from unfertilized oocytes through to the early developmental stage of the larvae.

제안 방법

KC442210 and AB052744, respectively) (Table 1). In order to investigate of olive flounder RAG-1 and IgM mRNA expression in various tissues, we assessed the expression of the both genes in various tissues from 90-days-old healthy olive flounder by RT-PCR (Fig. 1). It was found that RAG-1 and IgM mRNAs are significantly expressed in the spleen, kidney and gill, not expressed in other tissues.
However, the mRNA expression analysis of RAG-1 and IgM during the early developmental stage in olive flounder is insufficient. In this study, to information of RAG-1 and IgM expression during early larvae stages of development, we analyzed the expression pattern of both genes after hatching. As a result, RAG-1 and IgM mRNA was first detected in 5 dph, observed the increased mostly until 50 dph (Fig.
To investigate the expression of the RAG-1 and IgM genes, oligonucleotide specific primers were designed according to previously reported both genes in olive flounder (GenBank accession no. KC442210 and AB052744, respectively) (Table 1). In order to investigate of olive flounder RAG-1 and IgM mRNA expression in various tissues, we assessed the expression of the both genes in various tissues from 90-days-old healthy olive flounder by RT-PCR (Fig.

대상 데이터

Olive flounder larvae was obtained from Genetics and Breeding Research Center, National Fisheries Research and Development Institute (NFRDI; Geoje, Republic of Korea), and maintained in 10 tons flow through tank at 20±1℃ under a natural photoperiod. The sample was prepared from various tissues including brain, muscle, fin, eye, liver, gill, kidney, and spleen obtained from 90-day-old healthy olive flounder (n=3). Different larval stages including 5 to 55 days post hatching (dph) (5 dph, 10 dph, 15 dph, 20 dph, 25 dph, 30 dph, 35 dph, 40 dph, 45 dph, 50 dph and 55 dph) fish and unfertilized oocyte kept at 18.

데이터처리

The mRNA levels of RAG-1 and IgM was expressed as a ratio to those β-actin, which was simultaneously amplified as an internal control for each cDNA. The data was statistically analyzed by one-way ANOVA after arcsine transformation when needed, and followed by a Tukey test for identification of the statistically distinct groups. Significant differences were accepted for P <0.

성능/효과

1). The results confirmed in both genes that the spleen, kidney and gill are the typically immune organs in teleost fish species, since the differrentiation of early T and B cells occur in the tissues. In previous study, IgM gene was isolated and identified from the cDNA clones in olive flounder BAC library, and confirmed expression levels of various tissues using RT-PCR (Hirono et al, 2003).

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