Shekhar M S, Kiruthika J, Rajesh S, Ponniah A G
Genetics and Biotechnology Unit, Central Institute of Brackishwater Aquaculture, 75, Santhome High Road, R.A. Puram, Chennai, 600028, India,
Mol Biol Rep. 2014 Sep;41(9):6275-89. doi: 10.1007/s11033-014-3510-1. Epub 2014 Jun 29.
Four suppression subtractive hybridization (SSH) cDNA libraries were constructed to identify differentially expressed salinity stress responsive genes of black tiger shrimp, Penaeus monodon exposed to high (55 ppt) salinity conditions. One each of the forward and reverse SSH cDNA libraries were developed from the gill and gut tissues of shrimp and clones having inserts larger than 300 bp were unidirectionally sequenced. Based on the sequence homology search, the identified genes were categorized for their putative functions related to a wide range of biological roles, such as nucleic acid regulation and replication, immune response, energy and metabolism, signal transduction, cellular process, structural and membrane proteins, stress and osmoregulation. Gene expression levels in response to high salinity conditions at 2 weeks post salinity stress for some of the differentially expressed genes (Na(+)/K(+)-ATPase α-subunit, glutathione peroxidase, intracellular fatty acid binding protein, elongation factor 2, 14-3-3 like protein, penaeidin, translationally controlled tumor protein, transglutaminase and serine proteinase inhibitor B3) identified from SSH cDNA libraries were analysed by real-time RT-PCR. The highest gene expression levels was observed for Na(+)/K(+)-ATPase α-subunit in gill tissues (15.23-folds) and antennal glands (12.01-folds) and intracellular fatty acid binding protein in gut tissues (14.05-folds) respectively. The differential and significant levels of gene expression indicate the functional role of these genes in shrimp salinity stress adaptive mechanisms.
构建了四个抑制性消减杂交（SSH）cDNA文库，以鉴定在高盐度（55 ppt）条件下暴露的黑虎虾（斑节对虾）中差异表达的盐度应激反应基因。正向和反向SSH cDNA文库各一个分别从虾的鳃和肠道组织构建，对插入片段大于300 bp的克隆进行单向测序。基于序列同源性搜索，将鉴定出的基因根据其假定功能分类，这些功能与广泛的生物学作用相关，如核酸调控与复制、免疫反应、能量与代谢、信号转导、细胞过程、结构和膜蛋白、应激和渗透调节。对从SSH cDNA文库中鉴定出的一些差异表达基因（Na(+)/K(+)-ATP酶α亚基、谷胱甘肽过氧化物酶、细胞内脂肪酸结合蛋白、延伸因子2、14-3-3样蛋白、对虾抗菌肽、翻译控制肿瘤蛋白、转谷氨酰胺酶和丝氨酸蛋白酶抑制剂B3）在盐度应激后2周对高盐度条件的基因表达水平进行了实时RT-PCR分析。分别在鳃组织（15.23倍）和触角腺（12.01倍）中观察到Na(+)/K(+)-ATP酶α亚基的最高基因表达水平，在肠道组织中观察到细胞内脂肪酸结合蛋白的最高基因表达水平（14.05倍）。基因表达的差异和显著水平表明这些基因在虾盐度应激适应机制中的功能作用。
