Benchmark Agarose LE is a highly purified agarose, suitable for a variety of molecular biology applications. It is refined using an advanced process that excludes the use of organic solvents, yielding a cleaner end-product with a significantly reduced environmental impact. Agarose LE can be used for analyses of proteins and nucleic acids of various sizes (150 bp to 6 kbp). It's low EEO (= 0.13-mr ) promotes increased electrophoretic mobility, yielding improved resolution and shorter run times. This also allows macromolecules and larger particles (subcellular fragments, viruses, etc.) to migrate more freely through the gel matrix. The consistently low EEO also provides a reduction in band distortion (caused by counterflow) that can result from the presence of excessive sulfate-rich negative ions. Agarose LE is widely used for nucleic acid electrophoresis (analytical and preparative), protein electrophoresis (including radial immunodiffusion) and various blotting protocols. It is easily soluble, free of nucleases, and easy to use. It is highly transparent (forms a clear, colorless solution at 1g:100ml H2O), and exhibits exceptionally low absorption of chemical staining agents. Pore size can be adjusted by simple modifications to the concentration ratio. Formulated for high gel strength and integrity, Agarose LE exhibits exceptional thermal stability and mechanical resistance, ensuring safe, easy handling, regardless of whether a denaturing agent has been added. Gel Strength: >1200g/cm2 (1%)Gel Temperature: 360C + 1.50C (1.5%)Melt Temperature: 8800C + 1.50C (1.5%)EEO(-mr): 0.13Moisture Content: <7%Sulfate: <0.2%RNase/DNase: None DetectedProtease/Endonuclease: None DetectedStorage Conditions: Room Temperature
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1.怎么看两个反应-即目标基因和内参基因是否具有相同的扩增效率?
2.每次PCR反应的扩增情况都有差异,多次反应的扩增效率岂不是都不一样了吗?
3.如何设计才能让相对定量更可行?
由于对定量PCR的接触不多,希望各位高手多多帮助!谢谢!
所以想请大家给推荐一个效果好的最好是国产的质粒提取试剂盒。
多谢!
这些染料的结构式基本是保密的。有些地方能查到,象维基上就有SYBR Green的结果式,但谁也不能肯定是否是正确的。
1.熔融温度为75,74,73度的样品是否是同一个产物?熔融温度为56度的是不是没有扩增产物?
2.为什么相同的标准品不同反应CT值差别这么大?是不是荧光染料降解?我用的是试剂盒中的扩增酶,还没用几次。
3.扩增曲线的形态是否说明标准品已经有降解?
4.另外,我原来做CDNA梯度稀释做的挺好,可是这个直接是RNA为标准品,我用一步法进行QRT-PCR。标准品梯度稀释线性效果不稳定,有的时候好,有的时候不好。请假大家有没有好的建议?
附件中有扩增曲线和熔融曲线,请大家帮忙分析,谢过了!
新建MicrosoftOfficeWord文档.docx(46.18k)
既说像坐标画曲线曲线应纵轴该荧光强度吧轴数值变化代表荧光强度变化荧光强度变化代表钙离浓度变化 我觉直接用荧光强度值变化代替钙离浓度变化研究意义该钙离浓度变化吧
非要荧光强度值换算钙离浓度需要做列工作
1 取标准钙离浓度染色用共聚焦显微镜测荧光强度
2 标准品需要同像环境主要曝光间相同激光波及能量
3 标准品要用同浓度荧光染料
才能根据标准荧光强度关系推算品钙离浓度