| 袁 媛 陈 强 杨继红 熊国梅 刘中来.HBV X 区基因部分序列的亚克隆与PCR 检测[J].,2006,6(8):4-6 |
| HBV X 区基因部分序列的亚克隆与PCR 检测 |
| Subclone of partial sequence of gene of HBV X region and PCR detection |
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| DOI： |
| 中文关键词: 乙型肝炎病毒 X 区基因 亚克隆 灵敏度 |
| 英文关键词: HBV Gene of X region Sub- clone Sensitivity |
| 基金项目:国家自然科学基金资助项目( No. 40172005) |
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| 中文摘要: |
| 目的: 探讨HBV X 区基因用于乙型病毒性肝炎早期诊断价值和意义。方法: 设计特异性引物, 将pBR322- HBV 质粒
X 区部分序列PCR 产物AT 亚克隆至pBS- T 载体, 提取和纯化质粒DNA, 再对HBV X 区序列进行PCR 扩增。结果: 获得了预期
希望的质粒。PCR 的最佳退火温度为51 e , 灵敏度达到101 拷贝/ 2Ll, 线性范围101- 1010 拷贝/ 2Ll。讨论: pBR322- HBV 质粒
中的靶基因成功地被亚克隆至pBS- T 载体。pBR322- HBV 中X 区目的序列扩增产物为57bp, 该小片段勿需纯化就可直接AT
亚克隆至新载体, 有利于后续的常规PCR 检测和TaqMan MGB 荧光定量PCR 检测。 |
| 英文摘要: |
| Objective: To explore the significance and value of HBV X region genes in the early diagnosis of Hepatitis B. Methods: The
partial gene sequence of HBV X region in pBR322- HBV was amplified by PCR. The products amplified by PCR were sub- cloned into the pBS
- T vector, which were an effective AT clone plasmid. The positive AT sub- clones were sequenced. Results: The expected plasmid with HBV
X region gene was obtained. The optimal annealing temperature was 51 e . The sensitivity reached 101 copies per 2Ll. The linear rangewas 101
- 1010 copies per 2Ll. Discussion: The target genes in pBR322- HBV were sub- cloned to pBS- T. The amplification product of the target
gene sequence in pBR322- HBV was 57bp. It can be directly sub- cloned to the new vector without purification, which favors the routine PCR
and TaqManMGB fluorescent quantitative PCR for the early detection of HBV. |
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