Molecularhybridization分子杂交.pptVIP

Molecular hybridization  （分子杂交） 2. Probe (探针) Notes Probe is a specific DNA or RNA fragment which can bind with the sample DNA or RNA for detection. ATCCGATCG-------- Source of probe synthesized, cloning genomic DNA or cDNA, as well as RNA. Probe must be labeled(标记) before hybridization. radioactive αorγ32P-dCTP nonradioactive biotin, digoxigenin, fluorescent dye In a single stranded form for hybridization How a DNA probe bind with an unkown sequence 3. Ways of Molecular Hybridization A. Transfer blotting (转移印迹) Southern blotting Northern blotting Western blotting Eastern blotting? B. Dot blotting Slot blotting (点印迹, 狭缝印迹) C. In situ hybridization (原位杂交) A. Transfer blotting (转移印迹) Transfer blotting DNA or RNA is transferred and blotted from gel (胶)to membrane （膜）. Southern blotting It is first proposed by Dr. Edwin Southern in Edinburgh University in 1975, and term “Southern blotting” is named for him. Major steps: electrophoresis (电泳) transfer blotting （印迹转移） molecular hybridization（分子杂交） Three kinds of molecular hybridization Blotting Southern Northern Western Eastern Sample DNA RNA Protein ? Hybridization DNA-DNA RNA-DNA Protein-Protein molecule (Ag-Ab) 免疫印迹 B. Dot/Slot blotting (点印迹/狭缝印迹) Dot blot-DNA chip (芯片) Development and combination of multiple fields The specific array (阵列)of related genes. e.g., Oncogenes (癌基因) Tumor suppressor genes (抑癌基因) thousands,10 thousands of genes onto glass slide or silicon Characteristics high sensitivity (高灵敏), high throu