| PREPARE SOLUTIONS | |
| 1. SM buffer (1 L): | Mix 5.8 g of NaCl, 2 g of MgSO4-7H2O, 50 mL of 1M Tris-HCl, pH 7.5, 0.5 mL of 2% gelatin, and dH2O to 1 L (Autoclave) |
| 2. TENS buffer (100 mL): | Mix 5 mL of 1M Tris-HCl, pH 8.0 (50mM), 20 mL of 0.5 mM EDTA (100mM), 2 mL of 5M NaCl (100mM), 3 mL 10% SDS (0.3%), and 71 mL of dH2O |
| PROCEDURE | |
| 1. Plate cells and phage as described (144. cDNA library screening) and let plaques develop | |
| 2. Pippet enough SM buffer on plates to barely cover the plate and rock at 4oC for >1 hour | |
| 3. Pippet solution containing phage into a tube and add 0.2 mL of 2M ZnCl2 per 10 mL of SM solution | |
4. Centrifuge for 5 minutes at 5,000 rpms. A gray pellet forms | |
| 5. Resuspend pellet in TENS buffer | |
| 6. Heat at 65oC for 10 mins | |
| 7. Extract with phenol followed by chloroform:isoamyl alcohol | |
| 8. Precipitate with isopropanol:sodium acetate | |
9. Wash pellet with 70% ethanol | |
| 10. Resuspend pellet in T1E0.1, check DNA concentration and store aliquoted at -20oC |
RNA测序已经在生物学和医学的各个领域取得了前所未有的发展。在包括癌症在内的诸多疾病中，转录异构体的表达和用途是健康组织和患病组织之间变异的重要来源。鉴定差异剪接的异构体和融合转录本，可以为疾病的诊断......
造礁石珊瑚为珊瑚礁生态系统中的框架生物,研究其重要功能基因对于理解造礁石珊瑚对环境变化的响应有重要指示意义，近期来自中国科学院南海海洋研究所的研究人员提取了澄黄滨珊瑚(Poriteslutea)的总R......
Bio-Rad公司近日推出了一款“高级”的cDNA合成试剂盒，适用于定量PCR前的cDNA合成。此款名为iScriptAdvancedcDNAsynthesiskit的试剂盒能帮助研究人员生成更多的定......
今天在网上淘到的一本书GeneFlowfromGMPlants，05年，还算比较新。望大家多多支持。GeneFlowfromGMPlantsEditedbyGUYM.POPPYSchoolofBiol......
摘要:利用RT2PCR、RACE及克隆等方法获得了异育银鲫Toll样受体3(cagTLR3)基因全长cDNA序列。该cDNA全长3170bp,5′端非编码区181bp;3′端非编码区283bp;开放阅......
摘要:从革胡子鲶(Clariaslazera(Burchell))的脑垂体组织中提取总RNA,应用RT-PCR方法,扩增得到了革胡子鲶生长激素(Growthhormone,GH)基因cDNA的开放阅读......