| 欧阳胜荣 刘卓 唐认桥 李媛媛 赵欣 吴建新.NanoString nCounter 分析系统进行多基因表达计数的评价[J].,2015,15(3):413-416 |
| NanoString nCounter 分析系统进行多基因表达计数的评价 |
| Evaluation of multi-gene Expression Counting by NanoString nCounter |
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| DOI： |
| 中文关键词: NanoString nCounter 多基因表达计数 评价 |
| 英文关键词: NanoString nCounter Multi-gene expression counting Evaluation |
| 基金项目:国家重点基础发展规划项目（973项目）（2013CB530605） |
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| 中文摘要: |
| 目的：NanoString nCounter 是近年发展起来的进行基因表达谱检测的新平台，它利用分子条形码直接对基因表达进行多重
计数。本实验通过对mRNA 表达量的检测，分析nCounter 系统进行基因表达计数的可靠性。方法：应用NanoString 公司提供的
CAE Kit 试剂盒，通过杂交（mRNA样品与报告探针和捕获探针杂交）、纯化（清洗未结合探针）和计数（读取mRNA 的条数）三个
步骤，检测由人参考RNA 和人脑RNA构成的4 个样品中48个目的基因的mRNA 表达量；对获得的未进行标准化的原始数据，
应用Excel 2007 软件进行线性分析，从再现性、稳定性、准确性方面对nCounter分析系统进行评估。结果：再现性分析所有R2 均
大于0.998，线性拟合良好；试剂盒中阳性控制所有R2均大于0.99，高于质控要求的0.95；样品H70B30 和H30B70 的预期表达量
与实测值线性R2 大于0.999，预期值与实测值一致性良好。结论：nCounter 分析系统稳定性好，准确性高，检测mRNA 表达可
靠。 |
| 英文摘要: |
| Objective:The NanoString nCounter is a new platform for multi-gene expression counting using color- coded
molecular barcodes. The aim of this study was to evaluate the reliability of the nCounter Analysis System by detecting the mRNA
expression.Methods:The expression levels of 48 genes in 4 mRNA samples (composed of human references RNA and human brain
RNA) were detected using the CAE Kit provided by NanoString company. The procedure includes three steps of hybridization of mRNA(
reporter probes with capture probes), purification (washing off the uncombined probes) and mRNA counting. The unnormalized results
were evaluated by linear analysis using Excel 2007. Reproducibility, stability and accuracy of the nCounter Analysis System were
evaluated.Results:All of the R-squared value was larger than 0.998 by reproducibility analysis and the linear fitting was perfect.
R-squared value of the positive control was above 0.99, better than the specified data by quality control (0.95). R-squared value of the
expected values and actually measured values in H70B30 and H30B70 was larger than 0.999 by linear analysis, which indicated a
consistence of expected levels and actually measured values.Conclusion:nCounter Analysis System is stable, accurate and reliable on
the detection of mRNA expression. |
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