Heidenblut Anna M
Department of Internal Medicine, Knappschaftskrankenhaus. Ruhr-University, Bochum, Germany.
Methods Mol Biol. 2008;387:39-54. doi: 10.1007/978-1-59745-454-4_3.
In order to generate serial analysis of gene expression (SAGE) libraries from very small samples such as microdissected cells, the starting material must first be amplified via PCR or linear amplification of RNA. In microarray experiments, it has been shown that linear amplification of RNA can be used to generate reliable gene expression profiles and leads to the detection of expression differences that are not seen with nonamplified starting material. As the product of the amplification is amplified antisense RNA (aRNA), linear amplification of RNA cannot be used in combination with the conventional SAGE protocol. The aRNA-LongSAGE protocol described herein is an adaptation of the MicroSAGE protocol to the use of aRNA as starting material.
为了从诸如显微切割细胞等非常小的样本中生成基因表达系列分析(SAGE)文库,必须首先通过PCR或RNA的线性扩增对起始材料进行扩增。在微阵列实验中,已经表明RNA的线性扩增可用于生成可靠的基因表达谱,并导致检测到未扩增起始材料中未见的表达差异。由于扩增产物是扩增反义RNA(aRNA),因此RNA的线性扩增不能与传统的SAGE方案结合使用。本文所述的aRNA-LongSAGE方案是将MicroSAGE方案改编为使用aRNA作为起始材料。