关键词: 抑制消减杂交, 基因克隆, 差异表达基因, 反向mRNA 斑点印迹, 筛选

Abstract: Purpose : To establish a method for screening the differential expression genes from the suppression subt ractive hybridization positive clones. Methods : The PCR products of suppression subtractive hybridization positive clones were arrayed onto nylon membrane , hybridized with a labelled polyA + RNA probe and investi- gated with chemical luminescence reagent . The intensity of the dots was scanned and analyzed using Leica Q550 Iw image analysis system. The intensity of the dots were calculated as average intensity per unit area which eliminates the intensity of background. If the ratio (R) of intensity of a pair of dots in two parallel arrays is ≥ 2. 00 , it is defined to be positive differential expression gene. Results : The differential expression genes are efficaciously screened f rom suppression subtractive hybridization positive clones using reverse mRNA dot blot technique. Conclusion : The reverse mRNA dot blot is a useful method for screening differential expression genes from suppression subtractive hybridization positive clones.

Key words: suppression subtractive hybridization, gene clone, differential expression gene, reverse mRNA dot blot screen