酵母双杂交系统技术介绍.pptVIP

文库筛选: Mating方法筛选 Y187 α型 DDO筛选 二倍体 Y2H Gold a 型 Mating 接合反应 酵母的有性生殖过程， Clontech利用这个过程，实现了文库筛选。 Mix Mating QDO筛选 文库筛选: 均一化通用文库筛选 均一化通用型文库 高丰度与低丰度均衡 大大降低工作量 大大降低假阳性 更高的基因代表性 Normalized Universal Pre. library DDO/X/A 混合 QDO/X/A 文库筛选: 传统细菌文库筛选 文库培养 提取质粒 转化酵母菌 取1ml菌液按照一定比例用LB稀释，150ul/板进行涂板，过夜培养。 将提取的质粒转化Y187，获得酵母文库。 Mating筛选 将文库与诱饵培养液混合，进行Mating。涂筛选板，挑取阳性克隆子。 刮取平板上的菌落，提取质粒。 文库筛选: 验证阳性克隆子（PCR确认） 25ul 2×mix 30 PCR cycles 25ul H2O 630497 Matchmaker? Insert Check PCR Mix2 文库筛选: 酵母质粒提取 630467 Easy yeast Plasmid Isolation Kit 验证互作: 体内验证 SEAP 630305 Mammalian Matchmaker Two-Hybrid Assay Kit 2 验证互作: 体外Co-IP验证 Bait AcGFP1 PL Prey Protein A/G Agarose beads Matchmaker Chemiluminescent Co-IP Vector Set（630458） Matchmaker Chemiluminescent Co-IP Kit（630459） Prolabel Chemiluminescent Detection Kit （631723） 验证互作: 体外Co-IP验证实验流程 PL检测 验证互作: FRET分析 632466 pDsRed -Monomer-C1 Vector 632470 pAcGFP1 -C1-Vector Clontech Matchmaker系统产品选择指南 The FRIGIDA Complex Activates Transcription of FLC, a Strong Flowering Repressor in Arabidopsis, by Recruiting Chromatin Modification Factors K Choi, J Kim, HJ Hwang, S Kim… - The Plant Cell …, 2011 - Am Soc Plant Biol Extracellular signal-regulated kinase 8 (Erk8) controls estrogen-related receptor α (ERRα) cellular localization and inhibits its transcriptional activity M Rossi, D Colecchia, C Iavarone, A Strambi… - Journal of Biological …, 2011 – ASBMB WSB-1, a novel IL-21 receptor binding molecule, enhances the maturation of IL-21 receptor H Nara, T Onoda, M Rahman, A Araki… - Cellular …, 2011 – Elsevier FHL1 interacts with oestrogen receptors and regulates breast cancer cell growth L Ding, C Niu, Y Zheng, Z Xiong… - Journal of Cellular …, 2011 - Wiley Online Library Rab4 interacts with the human P‐glycoprotein and modulates its surface expression in multidrug resistant K562 cells C Ferrándiz‐Huertas… - … Journal of Cancer, 2011 - Wiley Online Library 6. Duplicated C-class MADS-box ge