Dual-Channel Surface Plasmon Resonance for Quantification of ApoE Gene and Genotype Discrimination in Unamplified Genomic DNA Extracts.
Dual-Channel Surface Plasmon Resonance for Quantification of ApoE Gene and Genotype Discrimination in Unamplified Genomic DNA Extracts.
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DOI：
10.1021/acssensors.8b00845
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发表时间：
2018-10
期刊：
影响因子：
8.9
通讯作者：
Xinyao Yi;Yonghong Xia;Binrong Ding;Ling Wu;Shengqiang Hu;Zixiao Wang;Minghui Yang;Jianxiu Wang
中科院分区：
文献类型：
--
作者：
Xinyao Yi;Yonghong Xia;Binrong Ding;Ling Wu;Shengqiang Hu;Zixiao Wang;Minghui Yang;Jianxiu Wang
基因变异的鉴定对于获得与疾病易感性相关的信息非常重要。建立了一种高灵敏度、高特异性的表面等离子体共振（SPR）定量检测载脂蛋白e基因的方法。与预固定的生物素化探针杂交后，具有GTGC碱基特异性识别位点的互补序列可被限制性内切酶HhaI切割，而单碱基错配（GTGC）的存在阻碍了切割反应的进行。在这两种情况下，链霉亲和素的掺入提高了SPR检测的灵敏度，对互补和单碱基错配序列的检测水平分别达到了10 fM和50 fM。传感方案简单，无标签，定量，从而避免了复杂的聚合酶链反应（PCR）扩增程序。提出的方法作为一个可行的手段，方便和敏感的分析载脂蛋白e基因在四个未扩增的基因组DNA提取物。
Identification of gene variation is of great importance for attaining information related to disease susceptibility. A highly sensitive and specific surface plasmon resonance (SPR) method for quantification of the apoE gene and genotype discrimination was demonstrated. The complementary sequences with the specific recognition sites of GCGC bases upon hybridization to the preimmobilized biotinylated probes could be cleaved by the restriction enzyme HhaI, while the existence of the single-base mismatch (GTGC) prevented the cleavage reaction. In both cases, the incorporation of streptavidin increased the sensitivity of the SPR assay, and the detection levels of 10 fM and 50 fM for the complementary and single-base mismatched sequences were attained, respectively. The sensing protocol is simple, label-free, and quantitative, thus avoiding the complicated polymerase chain reaction (PCR) amplification procedures. The proposed method serves as a viable means for facile and sensitive analyses of apoE genes in four unamplified genomic DNA extracts.