鲈鱼IgM重链基因cDNA的克隆及其原核表达

Abstract
Figure/Table
References
Related Citation (15)

Download: PDF (525 KB) HTML (1 KB)
Export: BibTeX | EndNote (RIS)

Abstract

In the present study, the complete cDNA sequence of the heavy chain of immunoglobulin M (IgM) in Lateolabrax japonicus was cloned using the method of homologous cloning and RACE technique. The full-length cDNA of IgM was 1,901 bp, and encoded a polypeptide of 582 amino acids. A phylogenetic tree was constructed based on the alignment of amino acid sequences from the constant regions in IgM heavy chain of L. japonicas and other related teleost fishes. The results showed that the IgM, IgZ, and IgD from various fishes clustered together, respectively, and formed three major branches, while the branches of IgM and IgZ generated from a single root suggested a closer evolutionary relationship between IgM and IgZ. RT-PCR results showed the highest IgM expression level in head-kidney, and an almost silence expression in heart, muscle and brain. The recombinant expression plasmid (pQE30-IgM) was constructed with encoding fragment of L. japonicas IgM heavy chain cDNA and the prokaryotic expression vector (pQE30), and transformed into Escherichia coli M15. The recombinant protein was identified by SDS-PAGE and Western blot using anti- 6×His monoclonal antibody which was specially binding with 6×His of the fusion protein.

Key words： Lateolabrax japonicus immunoglobulin M clone tissue distribution prokaryotic expression

Received: 25 April 2013

Q78

	Service

	E-mail this article
	Add to my bookshelf
	Add to citation manager
	E-mail Alert
	RSS
	Articles by authors
	LIU Yue
	ZHANG Qi-zhong

Cite this article:

LIU Yue,ZHANG Qi-zhong. Full-length cDNA cloning and prokaryotic expression of immunoglobulin M heavy chain in Weever (Lateolabrax japonicus)[J]. , 2013, 32(2): 218-223.

URL:

http://www.ecolsci.com/EN/ OR http://www.ecolsci.com/EN/Y2013/V32/I2/218

[1] 金伯泉. 医学免疫学[M]. 北京: 人民卫生出版社,2008. 34-36.
[2] Bang J D, Kim J W, Lee S D, Park S I, Chun S G, Jeong C S, Park J W. Humoral immune response of flounder to Edwardsiellatarda: the presence of various sizes of immunoglobulins in flounder[J]. Diseases Of Aquatic Organisms, 1996, 26: 197 -203.
[3] Kobayashi K.. Ontogenic Studies on the appearance of two classes of immunoglobulin forming cells in the spleen of the aleutian skate, Bathyraja Aleutica a Cartilayinous fish[J]. European journal of immunology, 1985, 15(9): 952-956.
[4] Saha N R, Suetake H, Kikuchi K, Suzuki Y. Fugu immunogloblin D: a highly unusual gene with unprecedented duplications in its constant region[J]. Immunogenetics, 2004, 56(6): 438-447.
[5] Danilova N, Bussmann J, Jekosch K, Steiner L A. The immunoglobulin heavy-chain locus in zebrafish: identification and expression of a previously unknown isotype, immunoglobulin Z[J]. Nature Immunology, 2005, 6(3): 295-302.
[6] Hansen J D, Landis E D, Phillips R B. Discovery of a unique Ig heavy-chain isotype (IgT) in rainbow trout: Implications for a distinctive B cell developmental pathway in teleost fish[J]. Proceedings of the National Academy of Science of the United States of America, 2005, 102(19): 6919-6924.
[7] Ram S, Azumi A, Miki N. Discovery of a novel immunoglobulin heavy chain gene chimera from common carp (Cyprinus carpio L.)[J]. Immunogenetics, 2005, 57(6): 458-463.
[8] Bernstein R M, Schluter S F, Shen S, Marchalonis J J. A new high molecular weight immunoglobulin class from the carcharhine shark: implications for the properties of the primordial immunoglobulin[J]. Proceedings of the National Academy of Science of the United States of America, 1996, 93(8): 3289-3293.
[9] Greenberg A S, Avila D, Hughes M, Hughes A, McKinney E C, Flajnik M F. A new antigen receptor family that undergoes rearrangement and extensive somatic diversification in sharks[J]. Nature, 1995, 374(6518): 168-173.
[10] Zhang Y A, Nie P, Wang Y P. cDNA sequence encoding immunoglobulin M heavy chain of the mandarin fish Siniperca chuatsi[J]. Fish & Shellfish Immunology, 2003, 14(5): 477-480.
[11] Iwama G, Nakanishi T. The fish immune system:organism, pathogen, and environment[M]. London: Academic Press, 1996.
[12] Cheng C A, John J A, Wu M S, Lee C Y, Lin C H, Lin C H, Chang C Y. Characterization of serum immunoglobulin M of grouper and cDNA cloning of its heavy chain[J]. Veterinary Immunology and Immunopathology, 2006, 109(3-4): 255-265.
[13] Mao M G, Lei J L, Alex P M. Characterization of RAG1 and IgM (mu chain) marking development of the immune system in red-spotted grouper (Epinephelus akaara)[J]. Fish & Shellfish Immunology, 2012, 33(4): 725-735.
[14] Nakao M, Moritomo T, Tomana M, Lee C Y, Lin C H, Lin C H, Chang C Y. Isolation of cdna encoding the constant region of the immunoglobulin heavy chain from common carp(Cyprirus carpio L.)[J]. Fish ＆ Shellfish Immunology, 1998, 8: 425-434.
[15] Srisapoome P, Ohira T, Hirono I, Aoki T. Genes of the constant regions of functional immunoglobulin heavy chain of Japanese flounder Paralichthys olivaceus[J]. Immunogenetics, 2004(4), 56: 292- 300.
[16] Feng J J, Guan R Z, Lin P, Guo S. Molecular cloning and characterization analysis of immunoglobulin M heavy chain gene in European eel (Anguilla anguilla)[J]. Veterinary Immunology and Immunopathology, 2009, 127(1-2): 144-147.
[17] 张福淼, 杨桂文, 李国田, 曹善东, 安利国. 鲈血清免疫球蛋白分离纯化及部分特性分析[J]. 动物学杂志, 2004, 39(5): 9-12.
[18] Cosciai M R, Morea V, Tramontano A, Oreste U. Analysis of a cDNA sequence encoding the immunoglobulin heavy chain of the Antarctic teleost Trematomus bernacchii[J]. Fish & Shellfish Immunology, 2000, 10(4): 343-357.
[19] 韩进刚, 付小哲, 石存斌, 李凯彬, 潘厚军, 吴淑勤. 剑尾鱼IgM基因的克隆及免疫对其组织表达的影响[J]. 广东海洋大学学报, 2007, 27(6): 1-6.
[20] Bause E. Structural requirements of n-glycosylation of proteins. Studies with proline peptides as conformational probes[J]. The Biochemical Journal, 1983, 209(2): 331-336.
[21] Lobb C J. Secretory immunity induced in catfish, Ictalurus punctatus, following bath immunization[J]. Developmental and Comparative Immunology, 1987, 11(4): 727-738.
[22] Grace M F, Manning M J. Histogenesis of the lymphoid organs in rainbow trout, Salmo gairdneri Rich[J]. Developmental and Comparative Immunology, 1980, 4(2):255-264.
[23] 李春涛. 黄颡鱼IgM分子及其表达研究[D]. 重庆:西南大学, 2012.
[24] 龚婷, 杨孝朴, 李银聚, 张春杰, 吴庭才, 程相朝. 鸡IL-2 全基因和去信号肽基因的原核表达[J]. 甘肃农业大学学报, 2009, 44(1): 11-14.