金属硫蛋白(MT),富含半胱氨酸的蛋白质的超家族,执行多个功能,例如维持必需金属的体内平衡,有毒金属的解毒和氧自由基的清除。在这项研究中,通过基因组DNA行走技术克隆了来自Bay扇贝Argopectenradians(设计为AiMT1)的金属硫蛋白(MT)基因的启动子区域,并对该区域的多态性进行筛选,以发现它们与对高温胁迫的易感性或耐受性的关联。在扩增的启动子区域中鉴定了一个插入缺失(ins-del)多态性和十六个单核苷酸多态性(SNP)。两个SNP,-375T-C和-337A-C,被选择来分析它们在从中国南部和北部沿海收集的两个海湾扇贝种群中的分布,被确定为耐热和易感库存,分别。有三种基因型,T/T,T/C和C/C,在基因座-375,它们的频率为25%,高温易感库存中的61.1%和13.9%,而34.2%,抗性股票中的42.1%和23.7%,分别。两种种群间不同基因型的频率分布差异无统计学意义(P>0.05)。相比之下,在基因座-337,三种基因型A/A,A/C和C/C显示频率为11.6%,热敏感股票中的34.9%和53.5%,而45.7%,耐热库存中的32.6%和21.7%,分别。C/C基因型在热敏感股票中的频率显著高于耐热股票(P<0.01),而耐热库存中A/A的频率明显高于易感库存(P<0.01)。此外,C/C基因型扇贝急性热疗120min后,AiMT1mRNA的表达明显高于A/A基因型(P<0.05)。这些结果表明,AiMT1位点-337的多态性与扇贝对热应激的易感性/耐受性有关,-337A/A基因型可能是未来选择具有耐热性的贝扇贝的潜在标记。
Metallothioneins (MTs), a superfamily of cysteine-rich proteins, perform multiple functions, such as maintaining homeostasis of essential metals, detoxification of toxic metals and scavenging of oxyradicals. In this study, the promoter region of a metallothionein (MT) gene from Bay scallop Argopecten irradians (designed as AiMT1) was cloned by the technique of genomic DNA walking, and the polymorphisms in this region were screened to find their association with susceptibility or tolerance to high temperature stress. One insert-deletion (ins-del) polymorphism and sixteen single nucleotide polymorphisms (SNPs) were identified in the amplified promoter region. Two SNPs, -375 T-C and -337 A-C, were selected to analyze their distribution in the two Bay scallop populations collected from southern and northern China coast, which were identified as heat resistant and heat susceptible stocks, respectively. There were three genotypes, T/T, T/C and C/C, at locus -375, and their frequencies were 25%, 61.1% and 13.9% in the heat susceptible stock, while 34.2%, 42.1% and 23.7% in the resistant stock, respectively. There was no significant difference in the frequency distribution of different genotypes between the two stocks (P>0.05). In contrast, at locus -337, three genotypes A/A, A/C and C/C were revealed with the frequencies of 11.6%, 34.9% and 53.5% in the heat susceptible stock, while 45.7%, 32.6% and 21.7% in the heat resistant stock, respectively. The frequency of C/C genotype in the heat susceptible stock was significantly higher (P<0.01) than that in the heat resistant stock, while the frequency of A/A in the heat resistant stock was significantly higher (P<0.01) than that in the heat susceptible stock. Furthermore, the expression of AiMT1 mRNA in scallops with C/C genotype was significantly higher than that with A/A genotype (P<0.05) after an acute heat treatment at 28°C for 120min. These results implied that the polymorphism at locus -337 of AiMT1 was associated with the susceptibility/tolerance of scallops to heat stress, and the -337 A/A genotype could be a potential marker available in future selection of Bay scallop with heat tolerance.