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目的运用基因芯片技术获取正常成人脑组织与人脑胶质瘤中差异表达的基因。方法抽提正常成人脑组织与人脑胶质瘤组织中的mRNA来制备探针,经杂交、洗涤后,通过计算机观察两者表达谱的差异情况,对507E08克隆子进行了Northern blot,生物信息学分析,蛋白表达和氨基酸测序。结果通过4次基因芯片筛选,获得15条与胶质瘤相关的新基因,经Northern blot 证实507E08基因在人正常脑组织中低表达,而在人脑胶质瘤中高表达。原位杂交得到相同的结果。BLASTn和BLAST分析显示,507E08基因为全长新基因,长度为2 002 bp,共编码203个氨基酸。本基因命名为人核糖体蛋白L14．22。经原核表达,在SDS-PAGE胶上获得一条22×103的条带,氨基酸序列分析显示,结果N-端15个氨基酸与序列所示的N-端15个氨基酸残基完全相同。结论基因芯片筛选正常脑组织与人脑胶质瘤差异表达的基因具有样品用量少,高质量,高速度, 高敏感等特性。507E08基因可能是与人脑胶质瘤形成有关的一条全长新基因。
Objective To use gene chip technology to obtain differentially expressed genes in normal adult human brain and human glioma. Methods The mRNA of normal adult brain tissue and human glioma tissue was extracted to prepare the probe. After hybridization and washing, the expression profile of the 507E08 clone was observed by computer. Northern blot, biological information Analysis, protein expression and amino acid sequencing. Results Fifteen novel genes related to glioma were obtained by four microarray analysis. Northern blot showed that 507E08 gene was low in normal human brain tissue and highly expressed in human glioma. In situ hybridization yielded the same result. BLASTn and BLAST analysis showed that the 507E08 gene was a full-length new gene with a length of 2 002 bp encoding a total of 203 amino acids. This gene is named human ribosomal protein L14.22. After prokaryotic expression, a 22 × 103 band was obtained on SDS-PAGE gel. Amino acid sequence analysis showed that the N-terminal 15 amino acids were exactly the same as the N-terminal 15 amino acid residues shown in the sequence. Conclusion Gene microarray screening of normal brain tissue and human glioma differentially expressed genes with less sample amount, high quality, high speed, high sensitivity and other characteristics. The 507E08 gene may be a new full-length gene associated with human glioma formation.