Xu Dong, Li Guangshan, Wu Liyou, Zhou Jizhong, Xu Ying
Protein Informatics Group, Life Sciences Division Environmental Sciences Division, Oak Ridge, TN 37831, USA.
Bioinformatics. 2002 Nov;18(11):1432-7. doi: 10.1093/bioinformatics/18.11.1432.
DNA microarray is a powerful high-throughput tool for studying gene function and regulatory networks. Due to the problem of potential cross hybridization, using full-length genes for microarray construction is not appropriate in some situations. A bioinformatic tool, PRIMEGENS, has recently been developed for the automatic design of PCR primers using DNA fragments that are specific to individual open reading frames (ORFs).
PRIMEGENS first carries out a BLAST search for each target ORF against all other ORFs of the genome to quickly identify possible homologous sequences. Then it performs optimal sequence alignment between the target ORF and each of its homologous ORFs using dynamic programming. PRIMEGENS uses the sequence alignments to select gene- specific fragments, and then feeds the fragments to the Primer3 program to design primer pairs for PCR amplification. PRIMEGENS can be run from the command line on Unix/Linux platforms as a stand-alone package or it can be used from a Web interface. The program runs efficiently, and it takes a few seconds per sequence on a typical workstation. PCR primers specific to individual ORFs from Shewanella oneidensis MR-1 and Deinococcus radiodurans R1 have been designed. The PCR amplification results indicate that this method is very efficient and reliable for designing specific probes for microarray analysis.
DNA微阵列是研究基因功能和调控网络的强大高通量工具。由于存在潜在交叉杂交问题，在某些情况下使用全长基因构建微阵列并不合适。最近开发了一种生物信息学工具PRIMEGENS，用于使用特定于各个开放阅读框（ORF）的DNA片段自动设计PCR引物。
PRIMEGENS首先针对基因组的所有其他ORF对每个目标ORF进行BLAST搜索，以快速识别可能的同源序列。然后，它使用动态规划在目标ORF与其每个同源ORF之间进行最佳序列比对。PRIMEGENS利用序列比对来选择基因特异性片段，然后将这些片段输入Primer3程序以设计用于PCR扩增的引物对。PRIMEGENS可以作为独立软件包在Unix/Linux平台的命令行上运行，也可以通过Web界面使用。该程序运行高效，在典型工作站上每个序列只需几秒钟。已经设计了针对希瓦氏菌MR-1和耐辐射球菌R1中各个ORF的PCR引物。PCR扩增结果表明，该方法对于设计用于微阵列分析的特异性探针非常有效且可靠。
