Colling Janine, Pollier Jacob, Makunga Nokwanda P, Goossens Alain
Department of Plant Systems Biology, VIB, Gent, Belgium.
Methods Mol Biol. 2013;1011:287-303. doi: 10.1007/978-1-62703-414-2_23.
cDNA-AFLP is a commonly used, robust, and reproducible tool for genome-wide expression analysis in any species, without requirement of prior sequence knowledge. Quantitative expression data are generated by gel-based visualization of cDNA-AFLP fingerprints obtained by selective PCR amplification of subsets of restriction fragments from a double-stranded cDNA template. Differences in gene expression levels across the samples are reflected in different band intensities on the high-resolution polyacrylamide gels. The differentially expressed genes can be identified by direct sequencing of re-amplified cDNA-AFLP tags purified from the gels. The cDNA-AFLP technique is especially useful for profiling of transcriptional responses of jasmonate-treated plants or plant (tissue) cultures and the discovery of jasmonate-responsive genes.
cDNA-AFLP是一种常用、可靠且可重复的工具,用于对任何物种进行全基因组表达分析,无需先验序列知识。通过对双链cDNA模板中限制性片段子集进行选择性PCR扩增获得cDNA-AFLP指纹图谱,基于凝胶的可视化产生定量表达数据。样品间基因表达水平的差异反映在高分辨率聚丙烯酰胺凝胶上不同的条带强度。通过对从凝胶中纯化的重新扩增的cDNA-AFLP标签进行直接测序,可以鉴定差异表达基因。cDNA-AFLP技术对于茉莉酸处理的植物或植物(组织)培养物的转录反应分析以及茉莉酸响应基因的发现特别有用。