2014, Volume 16, Issue 9
Strategic Study of CAE >> 2014, Volume 16, Issue 9
Full length cDNA cloning and expression of luteinizing hormone (LH) and which serum concentration was measured in half smooth tongue sole Cynoglossus semilaevis Günther
Qingdao Key Laboratory for Marine Fish Breeding and Biotechnology，Key Laboratory of Sustainable Development of Marine Fisheries，Ministry of Agriculture，Yellow Sea Fisheries Research Institute，Chinese Academy of Fishery Sciences，Qingdao 266071，China
Next Previous
Abstract
The full length cDNA of LH has been cloned from pituitary of half smooth tongue sole C. Günther by using RACE technology for the first time. The cDNA of LH is 670 bp，and the open reading frame is 477 bp encoding peptides of 158 amino acid. Multiple amino acid sequences alignments were performed among vertebrates，and the results showed that tongue sole LH mature peptide was the most similar to their orthologs of Pleuronectiformes and Perciformes which was 58 %～68 % identity. And 12 conserved cysteine residues and 1 N-linked glycosylation site were found in the deduced amino acid sequence. Real-time PCR analysis showed that LH mRNA was expressed in all twelve tissues including brain，pituitary，ovary and so on. The highest expression level was in pituitary. And the stronger amplification signals were detected in the brain and gonad than all other non- pituitary tissues. It suggested that LH may had the more physiological functions because LH mRNA of tongue sole expressed widely in non-pituitary tissues. Quantitative real- time PCR was used for analyzing the expression of female C. semilaevis LH mRNA in the reproductive cycle. The results showed that LH mRNA was expressed in all three tissues including brain，pituitary and ovary from the stage Ⅱto stage Ⅵ， and the highest expression level appeared on the stage Ⅳ and stage Ⅴwhich suggested that the main function of LH is to promote eggs finally mature and discharge. Radio immunoassay （RIA）marked by 125I was used to detect the concentration of plasma LH in C.semilaevis. The results showed that the biggest plasma LH concentration appareed on the stage of Ⅴwhich also suggested that the main function of LH was to promote eggs finally mature and discharge.
Keywords
Cynoglossus semilaevis Günther ; LH ; cDNA cloning ; Quantitative real-time PCR ; RIA
Figures
图1
图2
图3
图4
图5
图6
图7
图8
References
[ 1 ] Kajimura S，Yoshiura Y，Suzuki M，et al. cDNA cloning of two gonadotropin-β subunits（GTH-Ⅰβ and Ⅱ-β）and their expression profiles during gametogenesis in the Japanese flouder（Paralichthys olivaceus）[J]. General and Comparative Endocrinology， 2001，122（2）：117-129. link1
[ 2 ] Schulz R W，Viseher H F，Cavaco J E B，et al. GonadotroPins， their receptors，and the regulation of testicular functions in fish [J]. Comparative Biochemistry and Physiology Part B：Biochemistry and Molecular Biology，2001，129（2-3）：407-417. link1
[ 3 ] Schulz R W，Miura T. Spermatogenesis and its endocrine regulation [J]. Fish Physiology and Biochemisty，2002，26（1）：43-56. link1
[ 4 ] Swanson P，Dickey J T，Campbell B. Biochemistry and physiology of fish gonadotropins [J]. Fish Physiology and Biochemistry， 2003，28（1-4）：53-59. link1
[ 5 ] Prat F，Sumpter J P，Tyler C R. Validation of radioimmunoassay for two salmon gonadotropins（GTHⅠand GTHⅡ）and their plasma concentrations throughout the reproductivecycle in male and female rainbow trout（Oncorhynchus mykiss）[J]. Biology of Reproduction，1996，54（6）：1375-1382. link1
[ 6 ] 柳学周，庄志猛，马爱军，等. 半滑舌鳎繁殖生物学及繁育技术 研究[J]. 海洋水产研究，2005，26（3）：15-24. link1
[ 7 ] 柳学周. 半滑舌鳎繁殖及养殖技术（中）[J]. 科学养鱼，2006 （11）：14-15. link1
[ 8 ] 柳学周，庄志猛，马爱军，等. 半滑舌鳎苗种生产技术的开发研 究[J]. 海洋水产研究，2006，27（2）：17-24. link1
[ 9 ] 周 毅. 草鱼促性腺激素及其受体基因的克隆及表达分析[D]. 湖南：湖南师范大学，2009. link1
[10] Rosenfeld H，Levavi-Sivan B，Melamed P，et al. The GTH β subunit of tilapia：Gene cloning and expression [J]. Fish Physiology and Biochemistry，1997，1（1-6）：85-92. link1
[11] 张 勇，黄 海，李水生，等. 花鳗鲡促性腺激素cDNA克隆与 其在激素诱导性腺发育过程中的作用[R]. 2008.
[12] Kato Y，Gen K，Maruyama O，et al. Molecular cloning of cDNAs encoding two gonadotropin β subunits （GTH- Ⅰ βand GTH- Ⅱ β）from the masu salmon（Oncorhynchus masou）： Rapid divergence of the GTH-Ⅰβ gene [J]. Journal of Molecular Endocrinology，1993，11：275-282. link1
[13] 柳学周，孙中之，马爱军，等. 半滑舌鳎亲鱼培育及采卵技术的 研究[J]. 海洋水产研究，2006，27（2）：25-32. link1
[14] 柳学周，徐永江，刘乃真，等. 半滑舌鳎卵巢发育的组织学和形 态数量特征研究[J]. 渔业科学进展，2009，30（6）：25-35. link1
[15] Livak K J，Thomas D Schmittgen. Analysis of relative gene expression data using real-time quantitative PCR and the 2 − △ △Ct method [J]. Methods，2001，25（4）：402-408. link1
[16] Hearn M T，Gomme P T. Molecular architecture and biorecognition processes of the cystine knot protein superfamily：Part Ⅰ. The glycoprotein hormones [J]. Journal of Molecular Recognition，2000，13（5）：223-278. link1
[17] Power M E，Carolsfeld J，Wallis G P，et al. Isolation and characterization of a cDNA for gonadotropin Ⅱ- β of Pacific herring，an ancient teleost [J]. Journal of Fish Biology，1997，50： 315-323. link1
[18] Ulloa- Aguirre A，Timossi C，Damian- Matsumura P，et al. Role of glycosylation in function of follicle- stimulating hormone [J]. Endocrine，1999，11（3）：205-215. link1
[19] Parhar I S，Soga T，Ogawa S，et al. FSH and LH-β subunits in the preoptic nucleus：ontogenic expression in teleost [J]. Genral and Comparative Endocrinology，2003，132（3）：369-378. link1
[20] Wong T T，Zohar Y. Novel expression of gonadotropin subunit genes in oocytes of the gilthead seabream（Sparus aurata）[J]. Endocrinology，2004，145（11）：5210-5220. link1
[21] So W K，Kwok H F，Ge W. Zebrafish gonadotropins and their receptors：Ⅱ. Cloning and characterization of zebrafish folliclestimulating hormone and luteinizing hormone subunits - their spatial-temporal expression patterns and receptor specificity [J]. Biology of Reproduction，2005，72（6）：1382-1396. link1
[22] Natio N，Hyodo S，Okumoto N，et al. Differential production and regulation of gonadotropins（GTHⅠand GTHⅡ）in the pituitary gland of rainbow trout，Oncorhynchus mykiss，during ovarian development [J]. Cell and Tissue Research，1991，266 （3）：457-467. link1
[23] Prat F，Sumpter J P，Tyler C R. Validation of radioimmunoassay for two salmon gonadotropins（GTH Ⅰ and GTH Ⅱ）and their plasma concentrations throughout the reproductivecycle in male and female rainbow trout（Oncorhynchus mykiss）[J]. Biology of Reproduction，1996，54（6）：1375-1382. link1
[24] Gomez J M，Well C，Ollitrault M，et al. Growth hormone （GH）and gonadotropin subunit gene expression and pituitary and plasma changes during spermatogenesis and oogenesis in rainbow trout（Oncorhynchus mykiss）[J]. General and Comparative Endocrinology，1999，113（3）：413-428. link1
[25] Kim D J，Cho Y C，Sohn Y C. Molecular characterization of rockfish（Sebastes schlegeli）gonadotropin subunits and their mRNA expression profiles during oogenesis [J]. General and Comparative Endocrinology，2005，141：282-290. link1
[26] Kumar R S，Trant J M. Hypophyseal gene expression profiles of FSH-β，LH-β，and glycoprotein hormone-α subunits in Ictalurus punctatus throughout a reproductive cycle [J]. General and Comparative Endocrinology，2004，136：82-89. link1
[27] 陈晓燕. 促性腺激素受体在半滑舌鳎繁殖周期中的作用 研究[D]. 青岛：中国海洋大学，2010. link1
[28] Gomez J M，Well C，Ollitrault M，et al. Growth hormone （GH）and gonadotropin subunit gene expression and pituitary and plasma changes during spermatogenesis and oogenesis in rainbow trout（Oncorhynchus mykiss）[J]. General and Comparative Endocrinology，1999，113（3）：413-428. link1
[29] Trinh K Y，Wang N C，Hew C L，et al. Molecular cloning and sequencing of salmon gonadotropin β subunit [J]. European Journal of Biochemistry，1986，159（3）：619-624. link1
[30] Querat B，Hardy A，Fontaine Y A. Regulation of the type-Ⅱ gonadotropin α and β subunit mRNAs by estradiol and testosterone in the European eel [J]. Journal of Molecular Endocrinology，1991，7：81-86. link1
[31] Dickey J T，Swanson P. Effects of sex steroids on gonadotropin （FSH and LH） regulation in coho salmon（Oncorhynchus kisutch）[J]. Journal of Molecular Endocrinology，1998，21（3）： 291-306. link1
[32] Scott A P，Sumpter J P，Hardiman P A. Hormone changes during ovulation in the rainbow trout（Salmo gairdneri Richardson）[J]. Gen Comp Endocrinol，1983，49：128-134. link1
京公网安备 11010502051620号
