分子杂交-2016-4-11资料.pptVIP

Chips and Features 1.28 cm 1,300,000 features/chip Expression Assay Using Affymetrix GeneChip Now, we can infer which of the genes were expressed and in what intensity. Due to some biological processes, not always the correct sequence will hybridized to the oligo. The DNA Chip 蛋白质杂交 蛋白质三维结构域的契合 蛋白杂交：Westhern Blot Immunohistochemistry（ICH） Western blot 把经过电泳分离的蛋白组分从凝胶转移至一种固相支持体，以针对特定氨基酸序列的特异性试剂做为探针进行检测，通常使用的探针是抗体，它与附着于固相支持体的靶蛋白所呈现的抗原表位发生特异性反应。最后，结合上的抗体可用多种二级免疫学试剂检测。 Western blot Western blot p53 BNIP3 high metastasis potential lung cancer cells apoptosis Overexpression of Eukaryotic Initiation Factor 5A2Enhances Cell Motility and Promotes TumorMetastasis in Hepatocellular Carcinoma Dong-Jiang Tang，2010，Hepatology * Manufacturing Oligonucleotide Arrays Oligonucleotide arrays are produced in a way that is similar to the way computer chips are. We start with a matrix created over a glass substrate. Each cell in the matrix contains a ”chain” with appropriate chemical properties, and ending with an emterminator, a chemical gadget that prevents chain extension. This substrate is covered with a mask, covering some of the cells, but not others, and then illuminated. Covered cells are una.ected. In cells that are hit by the light, the bond with the terminator is severed. If we now expose the substrate to a solution containing a nucleotide base, it will form bonds with the non-terminated chains. Thus, some of the cells will now contain this nucleotide. The process can then be repeated with di.erent masks (which covers di.erent cells), and for di.erent nucleotides. This way one can insert a speci.c nucleotide to each cell of the matrix, and manufacture a speci.c oligonucleotide. Figure 1.16 demonstrates the production process. * ?????? ?? ???? ????? ???. ASO斑点杂交 正常探针 突变探针 PKU，AR 固相支持物 硝酸纤维素膜（nitrocellulose filter membrane) 较强吸附单链DNA和RNA的能力。 非特异性吸附蛋白能力较弱，适合蛋白方面的杂交 结合不牢、质地脆、不适于电转、小于200bp的DNA片段结合能力不强。 尼龙膜（nylon membrane) 单双链DNA和RNA结