Department of Physiology, Yeungnam University School of Medicine, Daegu 705-717, Korea.
Int J Mol Med. 2010 Jan;25(1):113-20.
In the present study, single-stranded large circular (LC)-sense molecules were utilized as probes for DNA microarrays and showed stronger binding signals than those of PCR-amplified cDNA probes. A microarray experiment using 284 LC-sense DNA probes found 6 upregulated and 7 downregulated genes in A549 cells as compared to WI38VA13 cells. Repeated experiments showed largely consistent results, and microarray data strongly correlated with data acquired from quantitative real-time RT-PCR. A large array comprising 5,079 LC-sense DNA was prepared, and analysis of the mean differential expression from dye-swap experiments revealed 332 upregulated and 509 downregulated genes in A549 cells compared to WI38VA13 cells. Subsequent functional analysis using an LC-antisense library of overexpressed genes identified 28 genes involved in A549 cell growth. These experiments demonstrated the proper features of LC-sense molecules as probe DNA for microarray and the potential utility of the combination of LC-sense and -antisense libraries for an effective functional validation of genes.
在本研究中，单链大环状 (LC)- 单链分子被用作 DNA 微阵列的探针，其与 PCR 扩增的 cDNA 探针相比显示出更强的结合信号。使用 284 个 LC- 单链 DNA 探针的微阵列实验发现，与 WI38VA13 细胞相比，A549 细胞中有 6 个上调和 7 个下调的基因。重复实验得到了大致一致的结果，并且微阵列数据与来自定量实时 RT-PCR 的数据高度相关。制备了包含 5079 个 LC- 单链 DNA 的大型阵列，并且对染料交换实验的平均差异表达分析显示，与 WI38VA13 细胞相比，A549 细胞中有 332 个上调和 509 个下调的基因。随后使用过表达基因的 LC- 反义文库进行的功能分析鉴定出 28 个参与 A549 细胞生长的基因。这些实验证明了 LC- 单链分子作为微阵列探针的适当特性，以及 LC- 单链和 - 反义文库组合用于有效验证基因功能的潜力。
