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넙치 (Paralichthys olivaceus) vasa 유전자의 full-length cDNA 분리 및 생식소 특이적 발현
정형복,김유철,김효원,Thanthrige Thiunuwan Priyathilaka,이성도,Viraj Udayantha Herath Mudiyanselage,최재영,황일선,진창남,허윤성,서종표,임봉수 제주대학교 해양과환경연구소 2014 해양과환경연구소 연구논문집 Vol.38 No.-
Until recent, primordial germ cells(PGCs) are recognized only by morphological observation, such as their large size and low nucleocytoplasmic ratio. For the molecular analysis of the reproduction, it is important to identify a specific marker of germ cell development and differentiation. The VASA, which was first identified in Drosophila, is reported as a germ-line cell specific marker gene in animals. Many other researches verified its germ cell specific expression during embryogenesis and gametogenesis. VASA is a member of the DEAD(Asp-Glu-Ala-Asp) protein family of ATP-dependent RNA helicase, and plays a critical role in germ-line cell linage. Vasa is expected to be an useful molecular marker for identification of PGCs in reproduction researches of aquaculture species. In this study, we isolated the vasa cDNA, and surveyed its gonad-specific expression in Paralichthys olivaceus. The full-length cDNA of P. olivaceus vasa cDNA was isolated and deduced amino acid sequence was compared to those of the other teleosts. It was 2,461bp long, consisted in 646 amino acids in its ORF region, 175bp of 5’-UTR, and 345bp 3’-UTR. Flounder VASA contained conserved DEAD box, arginine-glycine rich region, and other domains were found. Flounder vasa expressed strongly in the testis and ovary, confirming its property as an gonad-specific marker. These result is expected to be an useful marker for flounder reproduction research and related aquaculture industry.
노랑초파리 발생과정에서 sta 유전자의 발현 양상에 관한 연구
박지권,정형복,김세재 제주대학교 생명과학연구소 2000 제주생명과학연구 Vol.3 No.-
Drosophila stuvarista(sta) gene encodes the ribosomal protein D-p40, consisting of 270 amino acids. The D-p40 is highly conserved between such evolutionarily distinct organisms as human(63% identity) and hydra(58% identity) and has the effect of Minute phenotype such as other ribosomal proteins. We identified the sta gene in the process of Drosophila cDNA library screening and analyzed the developmental expression pattern of sta gene in the transcription level using in situ hybridization. Each animal stage and tissues of 3rd instar larva and adults were showed ubiquitously expression pattern. These results were similar to other ribosomal proteins that represented Minute phenotype. Also, we expressed the 31kDa recombinant proteins of D-p40 using his-tag expression system in E. coli and purified by affinity chromatography. Immunoblot result of the developmental stages was showedthat strongly detection only the stage of embryo and pupa. This result suggested that the expression of sta gene were regulated in the level of post-translation modification ailed considered that sta gene had to other function during development as well as to function as a core ribosomal component.
이성도,정형복,임봉수,이제희,김유철,김효원,THANTHRIGETHIUNUWAN PRIYATHILAKA,W. D. Niroshana Wickramaarachchi,김세재,김신권 한국유전학회 2014 Genes & Genomics Vol.36 No.4
Arginine vasotocin (AVT) is a neurohypophysialhormone of non-mammal vertebrates functions in variousphysiological processes and homologous to mammalvasopressin. In this study, a full-length cDNA coding for aputative AVT precursor was isolated from flounder, Paralichthysolivaceus by rapid amplification of cDNA ends. Itwas 641 bp long, coding 153 amino acids, and consisted in69 bp long 50-untranslated region (UTR), 462 bp openreading frame and 110 bp 30-UTR. The putative amino acidsequence of flounder AVT was determined, and comparedwith other AVTs from different species. Also, 1,447 bp offlounder AVT gene, corresponding to the isolated cDNAregion with 2 introns within it, was isolated. Reverse transcriptionpolymerase chain reaction result showed thatflounder AVT was strongly expressed in the brain but not inthe any other tissues examined, confirming it is brain-specificgene. Expression profile of the AVT, together with arylalkylamineN-acetyltransferase, heat shock protein 70,interferon-stimulated gene 15 and natural resistance-associatedmacrophage protein genes were surveyed fromStreptococcus iniae infected flounders by real-time quantitativePCR. It revealed that flounder AVT is up-regulatedunder acute stress compared to the non-challenged control.
현경만,정형복,임봉수,허성표,이영돈,박지권,김세재 한국유전학회 2010 Genes & Genomics Vol.32 No.6
In teleosts, gonadotropin-releasing hormone (GnRH) and gonadotropin hormone (GTH) play important roles in regulating gonadal development and maturation. In Southeast Asia, the longtooth grouper, Epinephelus bruneus, is a commercially important aquaculture fish. In this study, we cloned and characterized the longtooth grouper GnRH1 gene and cDNAs of three gonadotropin subunits (GTHα, FSHβ, LHβ). The GnRH1 gene of longtooth grouper was 4,032 bp long, and contained four exons, 61, 151, 99, and 423 bp long. GTHα, FSHβ, and LHβcDNAs were 509, 576, and 579 bp, respectively. Phylogenetic and Southern hybridization analyses revealed that the longtooth grouper GTH subunits were evolutionarily close to those of groupers and are one-copy genes. RT-PCR analyses showed that GTH subunit mRNAs were expressed at a higher level in the pituitary glands of immature fish than in those of mature fish, suggesting a role in gonadal maturation.
마정인,강선혜,정형복,이제희 한국수산과학회 2018 Fisheries and Aquatic Sciences Vol.21 No.1
Stimulator of interferon gene (STING) is induced by various inflammatory agents, such as lipopolysaccharide and microbial pathogens, including virus and bacteria. In this study, we obtained a full-length cDNA of a STING homolog from olive flounder using rapid amplification of cDNA ends PCR technique. The full-length cDNA of Paralichthys olivaceus STING (PoSTING) was 1442 bp in length and contained a 1209-bp open reading frame that translated into 402 amino acids. The theoretical molecular mass of the predicted protein sequence was 45.09 kDa. In the PoSTING protein, three transmembrane domains and the STING superfamily domain were identified as characteristic features. Quantitative real-time PCR revealed that PoSTING expressed in all the tissues analyzed, but showed the highest level in the spleen. Temporal expression analysis examined the significantly upregulated expression of PoSTING mRNA after viral hemorrhagic septicemia virus (VHSV) stimulation. In contrast, no significant changes in the PoSTING expression were detected in Edwardsiella tarda-challenged group compared to the un-injected control. The expression of P. olivaceus type I interferon (PoIFN-I) was also highly upregulated upon VHSV challenge. These results suggest that STING might be involved in the essential immune defense against viral infection together with the activation of IFN-I in olive flounder.
허성표,임봉수,황인준,김세재,유용운,허상우,송영보,정형복,백혜자,Akihiro Takemura,이영돈 한국통합생물학회 2012 Animal cells and systems Vol.16 No.2
We investigated the effects of fadrozol, an aromatase inhibitor (AI), and 17a-methyltestosterone (MT) on the induction of sex change in juvenile longtooth grouper Epinephelus bruneus, via histological observation of gonads. Changes in the mRNA expression of GtH subunits (FSH-b and LH-b) in the pituitary, and estradiol-17b (E2) and 11-ketotestosterone (11-KT) levels in the blood were also surveyed after AI and MT treatment. Juvenile longtooth groupers (113917 g body weight; 16.291.2 cm body length) received intramuscular injections of AI at 3 (3-AI) and 5 (5-AI) mg/kg BWdoses and MT at a 5 mg/kg BW (5-MT) dose. At week 7 post-injection, 3-AI and 5-MT oocytes were degenerated, and gonads of the 5-AI group initiated spermatogenesis. At week 21 post-injection, 3-AI- and 5-MT-treated gonads contained spermatogonia and spermatocytes, while 5-AI treatment induced advanced stages of spermatogenesis. The serum E2 level showed no significant differences throughout the experimental period, whereas that of 11-KT was significantly elevated in the 5-AI group at weeks 7 and 21 post-injection. A significant increase in the expression of FSH-b mRNA was evident in the 5-AI group at week 21 post-injection. In contrast, LH-b mRNA expression did not significantly differ among groups during the experimental period. These results imply that sex change has two stages in the longtooth grouper. In the first stage, oocytes are degenerated by the stimulation by 11-KT, and in the second stage spermatogenesis occurs, owing to the co-effects of 11-KT and FSH-b.