基于多环链接发夹型探针多聚荧光技术构建NSCLC耐药与循环miRNAs表达微阵列研究

The reports of accurate express chart between non-small cell cancers resistance drug and overall circle miRNAs could been found in world at present. The complicated amplify technologies such as stem-loop RT-PCR and RT-PCR with modified polyA are often used to detect the very short and little circle miRNA. The loop sequence of hairpin probe could detect miRNA because their lengths would been same. The hairpin probes immobilized on surface of materials could detect high-throughput miRNAs. However, it is difficult to immobilize hairpin probe on surface of materials. The hairpin probe with connected by two ends invented in the last project is important improve to solve the immobilization problem that could keep the "FRET". However, there is a technology limit of sensitivity and detectability. There are many previous study basements. We abandoned the traditional things and technology limit that single probe could been immobilized on single site, design and invent the new hairpin probe connected with many hairpins by change the stem structure. The many free hairpin probes connected with many hairpins could been gathered on the surface of nanoparticles, then the fluorescent signal could been gathered as many times. Therefore, the detectability and sensitivity could been improved as many times. The destination that highly sensitive and direct detecting the very little circle miRNAs without amplification could been achieved. It is important to NSCLC personalized medicine that building the microarray of the new hairpin probe connected with many hairpins to detect miRNAs about NSCLC resistance drug, and accurate express miRNAs chart.

非小细胞肺癌（NSCLC）化疗耐药与循环miRNAs精确表达谱国内外未见报道，明确miRNA表达与NSCLC耐药可减少用药盲目性。短小而微量的循环miRNA常需通过茎-环和加尾RT-PCR等复杂扩增进行检测。发夹探针环序列与miRNA长度相当而适用其检测；探针固定在载体表面可高通量检测miRNA，但受固定难题制约。断接式发夹探针是解决保留原始荧光FRET现象探针固定难题的重要进步，但存在灵敏度和检测限的技术瓶颈。基于前期研究基础，本研究打破载体表面单位点只能固定单个探针的局限性，以茎结构改造为突破口，设计了新型多环链接发夹探针：通过纳磁微球表面多个游离探针茎链接聚集和外磁场富集作用（即多聚荧光技术），显著提高灵敏度、拓宽检测限，并构建NSCLC耐药miRNA表达发夹探针微阵列及明确miRNA耐药表达谱，达到无需扩增/超灵敏检测循环miRNA的目的。该研究对NSCLC个体化用药等具重要意义。

非小细胞肺癌（NSCLC）化疗耐药与循环miRNAs精确表达谱国内外未见报道，明确miRNA表达与NSCLC耐药可减少用药盲目性。短小而微量的循环miRNA常需通过茎-环和加尾RT-PCR等复杂扩增进行检测。发夹探针环序列与miRNA长度相当而适用其检测；探针固定在载体表面可高通量检测miRNA，但受固定难题制约。断接式发夹探针是解决保留原始荧光FRET现象探针固定难题的重要进步，但存在灵敏度和检测限的技术瓶颈。基于前期研究基础，本研究打破载体表面单位点只能固定单个探针的局限性，以茎结构改造为突破口，设计了新型多环链接发夹探针：通过纳磁微球表面多个游离探针茎链接聚集和外磁场富集作用（即多聚荧光技术），显著提高灵敏度、拓宽检测限，并构建NSCLC耐药miRNA表达发夹探针微阵列及明确miRNA耐药表达谱，达到无需扩增/超灵敏检测循环miRNA的目的。该研究对NSCLC个体化用药等具重要意义。

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