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目的设计和克隆所有EB病毒(EBV)已知和预计的编码基因作为cDNA探针用于构建EBV微阵列,以促进研究EBV在其相关疾病中的发病学作用。方法cDNA探针首先由Oligo6.0, Blast和Primer Primier软件设计和筛选全EBV基因组探针,它们的长度被定在300~600 mer并且具有高度特异性。从B95-8细胞和NPC组织的基因组DNA和RNA中,通过PCR和RT-PCR方法扩增这些探针,之后克隆入T/A克隆载体。所有的探针被测序鉴定。结果除LF1和LF3基因在B95-8细胞的EBV基因组中不存在外,其余85个基因片段(BWRF1基因有7个重复阅读框架)和两个EBERs基因被成功克隆。结论EBV cDNA探针的设计和克隆为制备EBV微阵列和进一步研究EBV基因组在其相关疾病中的作用奠定了基础。
Objective To design and clone all Epstein-Barr virus (EBV) known and predicted coding genes as cDNA probes for the construction of EBV microarrays to facilitate the study of the pathogenesis of EBV in its related diseases. Methods cDNA probes The whole EBV genome probes were designed and screened by Oligo6.0, Blast and Primer Primier software. Their length was set at 300 ~ 600 mer and was highly specific. These probes were amplified by PCR and RT-PCR methods from the genomic DNA and RNA of B95-8 cells and NPC tissues and then cloned into the T / A cloning vector. All probes were sequenced. Results The other 85 fragments (BWRF1 gene has 7 repeat reading frames) and two EBERs genes were successfully cloned except LF1 and LF3 genes were not found in the EBV genome of B95-8 cells. Conclusion The design and cloning of EBV cDNA probes laid the foundation for the preparation of EBV microarrays and further study of the role of EBV genomes in their related diseases.