质粒DNA标准物质的分析_生物技术.doc

摘要:首先将质粒DNA标准物质载体细胞复活及放大培养，然后采用无内毒素质粒大提试剂盒提取质粒DNA，采用紫外分光光度法，芯片电泳法及PCR扩增三种定性方法对所提质粒DNA样品进行纯度验证，最后采用紫外法和电感耦合等离子体发射光谱法（ICP-OES）对所提质粒DNA样品进行定值分析，确定其含量。

结论：紫外法检测所提质粒DNA样品稀释后A260nm处的吸光度值及A260nm与A280nm处的吸光度比值分别在0.1-1.0和1.8-1.9合格范围之间；芯片电泳检测所提质粒DNA样品的条带图谱清晰，出峰位置都在4888-4997bp之间并且无其他杂峰，重复性较好；PCR检测所提质粒DNA样品的扩增曲线在特异性引物的控制下，图谱都很清晰，峰分离较好，标准曲线呈现一定的线性关系，可信度R2为0.993。紫外法测定质粒DNA样品的含量为1.2985±0.06mg/g，RSD为4.85%。ICP-OES测定质粒DNA样品的含量为1.2894±0.026 mg/g，RSD为2.02%，两种方法测定结果吻合度较高，综合证明了所提质粒DNA样品的纯度较高。

关键词：质粒DNA   提取及检测    分析

Abstract:First, the cell with plasmid DNA standard material carriers cultured, and then plasmid DNA extracted by the kit without endotoxin, and purity was determined by UV spectrophotometry, chip electrophoresis and PCR amplification. Final valuation analysis on the proposed plasmid DNA samples to determine its content by using UV spectrophotometry and inductively coupled emission spectrometry method (ICP-OES).

　　　Conclusion: The absorbance value of A260nm and A260nm/A280nm are between the range from 0.1-1.0 and 1.8-1.9 by UV-method; in chip electrophoresis of plasmid DNA samples has a clear peak position at 4888-4997bp with good reproducibility and no other impure peaks; the plasmid DNA amplified well in the control of specific primers, the map is very clear and the standard curve is linear, the credibility of R2 of 0.993. UV result for the determination of the content of the plasmid DNA samples was 1.2985±0.06mg/g, RSD was 4.85%. ICP-OES result of the content of the plasmid DNA samples was 1.2894±0.026 mg/g，RSD was 2.02%. Two results consistent with each other show that the mentioned plasmid DNA sample has a high purity.

Keywords: plasmid DNA   extraction and detection   analysis.

　　　经过紫外法检测，所提质粒DNA样品的稀释测定A260nm处的吸光度值及A260nm与A280nm处的吸光度比值分别在0.1-1.0和1.8-1.9合格范围之间；经芯片电泳检测，所提质粒DNA样品的条带图谱清晰，出峰位置都在4888-4997bp之间并且无其他杂峰，重复性较好；经PCR检测，所提质粒DNA样品的扩增曲线在特异性引物的控制下，图谱都很清晰，峰分离较好，标准曲线呈现一定的线性关系，可信度R2为0.993，综合证明所提质粒DNA样品的纯度较高。

紫外法测定质粒DNA样品的含量为1.2985±0.06mg/g，RSD为4.85%。ICP-OES测定质粒DNA样品的含量为1.2894±0.026 mg/g，RSD为2.02%，比较两种定量方法，我们可以看出其吻合度较高，保证了定量质粒DNA样品含量的实验结果真实有效。