Identification of cold responsive genes in Pacific white shrimp (Litopenaeus vannamei) by suppression subtractive hybridization.
Identification of cold responsive genes in Pacific white shrimp (Litopenaeus vannamei) by suppression subtractive hybridization.
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DOI：
10.1016/j.gene.2015.09.045
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发表时间：
2016-01
期刊：
影响因子：
3.5
通讯作者：
Jinxia Peng;P. Wei;Xiuli Chen;D. Zeng;Xiaohan Chen
中科院分区：
文献类型：
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作者：
Jinxia Peng;P. Wei;Xiuli Chen;D. Zeng;Xiaohan Chen
南美白色对虾（Litopenaeus vannamei）是世界上养殖面积最大的对虾品种之一。尽管如此由于原产于热带，目前在亚热带地区饲养，低温胁迫是对其生长、生存和分布的最严重威胁之一。揭示L.利用抑制性消减杂交（suppression subtractive hybridization，SSH）技术，对凡纳滨对虾肝胰腺中的冷应答基因进行了筛选。瓦纳梅。构建正向和反向cDNA文库，然后进行斑点杂交、克隆、序列分析和实时定量PCR。这些方法鉴定了92个冷诱导和48个冷抑制EST，得到总共37个冷诱导和17个冷抑制重叠群。一些与胁迫反应或细胞防御相关的基因，如四转膜蛋白（tetraspanins，TSPAN）、DEAD-box解旋酶、热休克蛋白（heat shock proteins，HSPs）和金属硫蛋白（metallothionein，MT）在正向SSH文库中的表达量高于反向SSH文库。最丰富的Est是四跨膜蛋白-8（tetraspanin-8，TSPAN 8）同系物，称为LvTSPAN 8。对LvTSPAN 8基因进行了多序列比对和跨膜结构域预测，并检测了冷暴露后鳃、肌肉、心脏和肝胰腺中LvTSPAN 8的表达，结果表明LvTSPAN 8在肝胰腺中表达量最高。总体而言，这项研究能够通过SSH鉴定出几个已知的基因和新的基因，这些基因似乎与冷胁迫有关，这将有助于进一步了解L.瓦纳梅。
The Pacific white shrimp (Litopenaeus vannamei) is one of the most widely cultured shrimp species in the world. DespiteL. vannameihaving tropical origins, it is being reared subtropically, with low temperature stress being one of the most severe threats to its growth, survival and distribution. To unravel the molecular basis of cold tolerance inL. vannamei, the suppression subtractive hybridization (SSH) platform was employed to identify cold responsive genes in the hepatopancreas ofL. vannamei. Both forward and reverse cDNA libraries were constructed, followed by dot blot hybridization, cloning, sequence analysis and quantitative real-time PCR. These approaches identified 92 cold induced and 48 cold inhibited ESTs to give a total of 37 cold induced and 17 cold inhibited contigs. Some of the identified genes related to stress response or cell defense, such as tetraspanins (TSPANs),DEAD-box helicase, heat shock proteins (HSPs) and metallothionein (MT), which were more abundant in the forward SSH library than in the reverse SSH library. The most abundant Est was atetraspanin-8(TSPAN8) homolog dubbedLvTSPAN8. A multiple sequence alignment and transmembrane domain prediction was also performed forLvTSPAN8.LvTSPAN8 expression was also examined in the gills, muscle, heart and hepatopancreas following cold exposure and showed the highest expression levels in the hepatopancreas. Overall, this study was able to identify several known genes and novel genes via SSH that appear to be associated with cold stress and will help to provide further insights into the molecular mechanisms regulating cold tolerance inL. vannamei.