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为了克隆与白血病复发相关的新基因 (LRP15 )的全长cDNA序列 ,应用分子生物学及生物信息学技术对一段已知仅在白血病复发标本中被甲基化的 1.8kb长的DNA片段进行研究分析 ,通过美国生物信息中心 (NCBI)提供的hEST数据库进行电子杂交 ,并对获得的相互重叠的EST片段进行组装 ,设计引物进行cDNA末端快速扩增(RACE)。以高通量基因组序列 (HTGS)数据库及SAGE文库为基础 ,将获得的cDNA片段进行染色体定位及组织表达分析。结果表明 ,LRP15基因的cDNA序列全长 1718bp ,含 1个 780bp的开放读码框架 ,编码 2 5 9个氨基酸。该基因定位于染色体 3p2 4 ,在多种组织中表达。结论 :RACE技术是克隆新基因的有效方法 ,LRP15可能是一个与细胞恶变及白血病复发相关的候选基因
In order to clone the full-length cDNA sequence of a novel gene (LRP15) associated with relapse of leukemia, a 1.8 kb DNA fragment known to be methylated only in leukemia recurrent specimens was studied using molecular biology and bioinformatics techniques After analysis, the hybridization was performed by the hEST database provided by the American Center for Biological Information (NCBI), and the obtained overlapping EST fragments were assembled. Primers were designed for rapid amplification of cDNA ends (RACE). Based on the high-throughput genomic sequence (HTGS) database and the SAGE library, the obtained cDNA fragments were subjected to chromosome mapping and tissue expression analysis. The results showed that the cDNA sequence of LRP15 gene was 1718bp in length and contained a 780bp open reading frame encoding 259 amino acids. This gene is located on chromosome 3p24 and is expressed in many tissues. Conclusion: RACE technique is an effective method for cloning new genes. LRP15 may be a candidate gene associated with cell malignant transformation and leukemia recurrence