应用微阵列技术筛选HBV前-S1蛋白的反式调节基因谱

目的 阐明前-S1蛋白的表达对乙型肝炎病毒(HBV)感染肝细胞基因表达谱的影响。方法 以含有HBV全基因组的质粒G376-7(GenBank号：AF384371)作为模板，应用PCR扩增前-S1蛋白编码基因片段，以常规的分子生物学技术构建表达载体pcDNA3．1(-)-preS1，以脂质体转染肝母细胞瘤细胞系HepG2，提取总mRNA，逆转录为cDNA，与转染空白表达载体pcDNA3．1(-)的HepG2细胞进行DNA芯片分析并比较。结果 在1152个基因表达谱的筛选中，发现有30个基因表达水平显著上调，38个基因表达水平显著下调。结论 前-S1基因的表达对于肝细胞基因表达谱有显著影响。DNA芯片技术是分析反式调节靶基因的有效技术途径。

Screening and identification of the genes trans-regulated by hepatitis B virus pre-S1 protein with microarray assay

Objective To evaluate the influence of expression of pre S1 protein on the genomic expression of hepatitis B virus (HBV) infected hepatocyte with microarray. Methods The differentially expressed genes between the hepatoblastoma cell line HepG2 transfected by pcDNA3.1(-) and pcDNA3.1(-)-preS1, were respectively compared by cDNA microarray technique. The HBV pre-S1 coding DNA fragment was amplified with polymerase chain reaction (PCR) technique by using G376-7 plasmid DNA containing the full length of HBV genome as the template. The expressive vector of pcDNA3.1-preS1 was constructed by routine molecular biological methods. HepG2 cells were transfected by pcDNA3.1(-) and pcDNA3.1-preS1, respectively, using FuGENE6 Transfection Reagent. The total RNA was isolated and reversely transcribed. Results The cDNAs were subjected for microarray screening with 1152 cDNA probes. From the scanning results, it was found that 30 genes were up-regulated and 38 genes were down-regulated by pre-S1 protein of HBV. Conclusion The expression of pre-S1 protein affected the genomic expression spectrum of HBV infected hepatocyte(HepG2 cell line).