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溶菌酶是机体先天免疫系统中一个重要的效应分子,参与机体多种免疫反应。本研究克隆了近江牡蛎Crassostrea hongkongensis i-型溶菌酶的保守cDNA序列,并探讨其在不同组织的表达、不同温度对其表达的影响。结果表明,近江牡蛎i-型溶菌酶保守cDNA序列长634 bp,包含一个长420 bp开放阅读框,编码139个氨基酸;其蛋白序列具有典型的i-型溶菌酶特征,包含特定氨基酸残基序列CL(E/L/R/H)C(I/M)C、部分高度保守序列SCG(P/Y)FQI和酶活性中心位点(Glu34、Asp45、Ser48、Trp61);另外,还发现一个新的氨基酸保守序列HNGGPRGC。i-型溶菌酶基因在近江牡蛎消化腺、肌肉、鳃、外套膜、唇瓣等5种组织的表达量以消化腺最高,鳃次之,肌肉最低。i-型溶菌酶基因的表达与温度相关,水温20—25℃,其表达量比较稳定;水温6℃或13℃,表达量都呈下降趋势;在27℃,表达量在整个实验过程都维持一个相对高的水平;在34℃,其表达量在第12小时达到最高,第48小时后急剧下降;表明温度可影响i-型溶菌酶基因表达,从而调节其免疫功能和对外环境的应激反应。
Lysozyme is an important effector molecule in the body’s innate immune system and participates in a variety of immune responses in the body. In this study, we cloned the conserved cDNA sequence of Crassostrea hongkongensis i-type lysozyme in Oyster oyster and explored its expression in different tissues and different temperature. The results showed that the conserved cDNA sequence of i-type lysozyme was 634 bp in length and contained a 420 bp open reading frame encoding 139 amino acids. The protein sequence of i-type lysozyme was characterized by typical i-type lysozyme, including specific amino acid residue sequence (P / Y) FQI and enzyme active site (Glu34, Asp45, Ser48, Trp61) in addition to the C (I / M) C and some highly conserved sequences The new amino acid conserved sequence HNGGPRGC. The expression level of i-type lysozyme gene in the digestive gland, muscle, gill, mantle and labium of Oyster oyster was the highest in the digestive gland, followed by the gill followed by the lowest muscle. The expression of i-type lysozyme gene was correlated with temperature. The water temperature was 20-25 ℃. The expression level of i-type lysozyme gene was stable. When the water temperature was 6 ℃ or 13 ℃, the expression level decreased. At 27 ℃, the expression level was maintained throughout the experiment A relatively high level; at 34 ℃, its expression reached the highest at the 12th hour, the sharp decline after 48 hours; that the temperature can affect the i-type lysozyme gene expression, thereby regulating its immune function and external environment stress reaction.