黄鳝csf1r基因的克隆及时空表达特征分析

【目的】克隆黄鳝csf1r基因，并对其时空表达特性进行分析，为探明csf1r基因在黄鳝不同体色形成中的作用奠定基础。【方法】采用RACEs（Rapid-amplification of cDNA ends）技术从黄鳝皮肤cDNAs中克隆得到csf1r基因的全长cDNA序列，对其编码蛋白进行生物信息学分析。采用实时荧光定量PCR（qRT-PCR）方法检测csf1r基因在黄鳝不同组织、不同发育时期胚胎或个体及3种体色黄鳝（黄黑斑鳝、碎花斑鳝和隐花斑鳝）皮肤和肾脏中的相对表达量，分析该基因的表达特征。测定3种体色黄鳝肝脏中的碱性磷酸酶（AKP）、超氧化物歧化酶（SOD）活性和总抗氧化能力（T-AOC）。【结果】黄鳝csf1r cDNA序列全长为4 430 bp（GenBank收录号：OP589303），其编码区长度为2 937 bp，编码978个氨基酸，存在免疫球蛋白结构域和蛋白激酶催化结构域2个保守结构域。荧光定量PCR结果表明，csf1r基因在黄鳝脑、精巢、卵巢、肠、心脏、肾脏、肝脏、肌肉、皮肤和脾脏等组织中均有表达，在脾脏和心脏中表达量较高，其次是肾脏、皮肤和肌肉，卵巢中表达量最低；csf1r在胚胎眼晶体形成期开始大量表达，显微观察发现该时期胚胎的躯干上开始有色素颗粒出现。在3种体色黄鳝皮肤和肾脏中，csf1r基因在黄黑斑鳝的皮肤中表达量最低，而在其肾脏中表达量最高。3种体色黄鳝肝脏氧化应激指标测定结果发现，黄黑斑鳝肝脏中的碱性磷酸酶（AKP）、超氧化物歧化酶（SOD）活性和总抗氧化能力比其他2种体色黄鳝高，但是差异未达显著水平。【结论】csf1r基因可能不仅参与了黄鳝体色的形成，还与黄鳝非特异性免疫相关。

Cloning and spatio-temporal expression pattern of csf1r gene in Monopterus albus

【Objective】The colony-stimulating factor 1 receptor (csf1r) gene of Chinese swamp eel (Monopterus albus) was cloned and its spatio-temporal expression pattern was profiled to provide a basis for further exploring roles of csf1r gene in body color formation of Monopterus albus.【Method】The cDNA sequence of csf1r gene of Chinese swamp eel was cloned from the cDNA of skin by Rapid-amplification of cDNA ends (RACEs),and the deduced amino acid sequence was analyzed.Expression levels of csf1r gene were detected in different tissues,different developmental stages of embryos or individual and tissues of skin and kidney of swamp eels with three body colors (black-yellow brinded Monopterus albus,small-brinded and hidden-brinded Monopterus albus).The activities of alkaline phosphatase (AKP),superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) in liver of Monopterus albus were also determined.【Result】The full-length cDNA of csf1r was 4 430 bp in length (GenBank accession number:OP589303),and the open reading frame was 2 937 bp,encoding 978 amino acids.It included two conserved domains of immunoglobulin domain and protein kinase catalytic domain.PCR results showed that csf1r expressed in all tissues with the highest in spleen and heart,followed by kidney,skin and muscle,and the least in ovary.The expression of csf1r was firstly detected during the stage of lens formation,and pigment granules were present in the embryo torso.The expression level of csf1r gene was the lowest in skin and the highest in kidney of black-yellow brinded Monopterus albus among three varieties.Activities of AKP,SOD and T-AOC in black-yellow brinded Monopterus albus were higher than in others.【Conclusion】The csf1r gene might be involved in body color formation and nonspecific immunity.