淡水养殖凡纳滨对虾生长相关基因HDAC1克隆及其表达分析

Cloning and expression analysis of HDAC1 gene related to growth of Litopenaeus vannamei cultured in freshwater

【目的】分析组蛋白去乙酰基酶1基因(HDAC1)在不同淡水生长特性家系凡纳滨对虾中的表达情况,明确HDAC1基因在对虾淡水养殖过程中的功能作用,为揭示凡纳滨对虾生长调控的分子机制提供参考依据。【方法】通过RACE克隆凡纳滨对虾HADC1基因cDNA序列,利用DNAMAN、ProtParam、ProtScale、PSORT等在线软件进行生物信息学分析,以实时荧光定量PCR检测凡纳滨对虾HDAC1基因组织表达特征;选取4个家系凡纳滨对虾在1‰盐度下进行淡水养殖,养殖70 d后统计对虾存活率(RS)、日增重率、体长日增率、特定生长率(SGR)及饲料转化率(FCR),并利用实时荧光定量PCR检测凡纳滨对虾肠道中的HDAC1基因表达情况。【结果】凡纳滨对虾HDAC1基因c DNA全长3230 bp,包括67 bp的5’非编码区(5’-UTR)、1711 bp的3’非编码区(3’-UTR)及1452 bp的开放阅读框(ORF),共编码483个氨基酸残基;HDAC1基因DNA全长4692 bp,含有8个外显子和7个内含子。凡纳滨对虾HDAC1基因编码蛋白分子量为54640.22 Da,理论等电点(p I)为5.59,属于稳定的亲水性蛋白,具有1个Histone deacetylase 1结构域,无跨膜结构。凡纳滨对虾HDAC1氨基酸序列与斑节对虾HDAC1氨基酸序列的相似性最高,达99.79%。HDAC1基因在凡纳滨对虾各组织中均有表达,以在肠道中的相对表达量显著高于在其他组织中的相对表达量(P<0.05);在4个家系群体中,无论是大虾还是小虾,生长慢的群体中HDAC1基因相对表达量较低,生长快的群体中HDAC1基因相对表达量较高。【结论】HDAC1基因在凡纳滨对虾各组织中均有表达,以肠道中的相对表达量最高,且HDAC1基因相对表达量与凡纳滨对虾的生长速度成正比,故推测其参与凡纳滨对虾的生长调节过程。

【Objective】To analyze the expression of histone deacetylase 1 gene(HDAC1)in shrimp families with different freshwater growth characteristics,and to clarify the function of HDAC1 gene related to growth of Litopenaeus vannamei cultured in freshwater,so as to provide a reference for revealing the molecular mechanism of growth regulation of L.vannamei.【Method】The cDNA sequence of HDAC1 gene of L.vannamei was cloned by RACE,and bioinformatics analysis was carried out by using online software such as DNAMAN,ProtParam,Protscale and PSORT.The tissue expression characteristics of HDAC1 gene of L.vannamei were detected by real-time fluorescent quantitative PCR.Four families of L.vannamei were cultured in fresh water at 1‰salinity.After 70 days of culture,the survival rate(RS),daily gain rate,daily gain rate of body length,specific growth rate(SGR)and feed conversion rate(FCR)of L.vannamei were counted,and the expression of HDAC1 gene in the intestine of L.vannamei was detected by real-time fluorescent quantitative PCR.【Result】The full length of HDAC1 gene cDNA of L.vannamei was 3230 bp,including 67 bp 5’-UTR,1711 bp 3’-UTR and 1452 bp open reading frame(ORF),encoding 483 amino acids in total.The total length of HDAC1gene DNA was 4692 bp,containing eight exons and seven introns.The HDAC1 gene of L.vannamei encoded a protein with a molecular weight of 54640.22 Da and a theoretical isoelectric point of 5.59,and it was a stable hydrophilic protein with a histone deacetylase 1 domain with no transmembrane structure.The similarity of HDAC1 amino acid sequence between L.vannamei and Penaeus monodon was the highest,up to 99.79%.HDAC1 gene was expressed in all tissues of L.vannamei,and the relative expression in intestine was significantly higher than that in other tissues(P<0.05).Among the four pedigree populations,whether big shrimp or small shrimp,the relative expression of HDAC1 gene was lower in the slow-growing population,and higher in the fast-growing population.【Conclusion】HDAC1 gene is expressed in all tissues of L.vannamei,with the highest relative expression in the intestine,and the relative expression of HDAC1 gene is directly proportional to the growth rate of L.vannamei,so it is speculated that it is involved in the growth regulation process of L.vannamei.