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目的 探讨寡核苷酸芯片检测点突变过程中 ,不同类型探针在预先化学处理的玻片表面上固定效率的不同对于杂交信号的影响。方法 根据HLA DRB1 12亚型的基因序列设计四种 5′末端不同化学基团修饰的探针 ,通过与醛基化玻片和溴化玻片结合形成两种寡核苷酸微阵列芯片 ,与不对称荧光标记的目的片段杂交 ,通过荧光信号的强度判断最佳类型探针。在此基础上进一步设计两组四种碱基突变探针 ,根据杂交结果 ,判断最佳检测突变探针。结果 5′末端氨基探针和硫代探针可以分别固定在两种类型的玻片上 ,溴化芯片中杂交信号强于醛基化芯片的杂交信号。其中 5′末端带有连接臂的氨基探针在检测点突变中阳性信号同阴性信号比值要高于 5′末端硫代探针 ,重复实验判型结果稳定一致。结论 溴化芯片中 5′末端带有连接臂的氨基探针在检测点突变中具有更明显的优势 ,为进一步的研究和制备点突变寡核苷酸芯片提供了条件
OBJECTIVE: To investigate the effect of different immobilization efficiencies of different types of probes on the surface of pre-chemically treated glass slides during the detection of point mutations in oligonucleotide chips. Methods Based on the gene sequence of HLA DRB1 12 subtype, four probes with different chemical groups at the 5 ’end were designed and used to form two oligonucleotide microarray chips by combining with aldehyde-based glass slides and bromide glass slides. Asymmetric fluorescently labeled target fragments hybridize and the best type of probe is judged by the intensity of the fluorescent signal. Based on this, two groups of four base mutation probes are further designed, and the best detection mutation probe is judged according to the hybridization result. As a result, the 5 ’terminal amino probe and the thio probe can be respectively immobilized on two types of slides, and the hybridization signal in the brominated chip is stronger than that of the aldehyde-based chip. Among them, the amino probe with the linker at the 5 ’end was higher than the negative signal at the detection point mutation, which was consistent with that of the 5’ end probe. Conclusion Amino-probe with a linker at the 5 ’end of the brominated chip has more obvious advantages in detection of point mutation, which provides the conditions for further research and preparation of point mutation oligonucleotide chips