摘要
目的 应用现有DNA序列分析技术对人类免疫缺陷病毒 1型 (HIV 1)的蛋白酶 (PR)和逆转录酶 (RT)基因进行简便、快速、准确的序列分析 ,在临床上为HIV 1患者治疗方案的确定提供可靠依据。方法 从HIV 1感染者血清中提取RNA ,用套式逆转录 聚合酶链反应 (RT PCR)扩增HIV 1的PR和RT基因片段 ,并对此片段进行核苷酸序列分析。结果 本方法对血清中HIV 1RNA含量约 2 0 0 0拷贝 ml以上的样品都能有效扩增。在序列分析时背景干扰少 ,对 2 5 %左右的混合序列均能准确反映。通过对临床治疗病人进行动态序列分析观察 ,发现在药物治疗过程中 ,病人血浆中HIV 1RNA含量变化与PR和RT基因抗药性变异具有很大相关性。结论 PR和RT基因序列分析方法能够及时、准确地反映HIV 1患者在抗病毒药物治疗过程中病毒的变异情况。该方法的建立能在临床上能为不同患者治疗方案的确定提供直接。
Objective To develop and validate a rapid and sensitive method that can be used to detect emergence of HIV 1 drug resistant viruses in HIV infected patients receiving highly active antiretroviral therapies (HAART). Methods HIV 1 viral RNA was extracted from plasma and amplified by nested RT PCR. Nucleotide sequences of the reverse transcriptase (RT) and protease (PR) genes were determined by automated DNA sequencing. Results RT and PR sequences can be reliably determined with a minimum 2 000 copies/ml of viral RNA in plasma. Emergence of drug resistant viruses can be detected as soon as it reached to >25% of the total viral populations. By using this method, drug resistant viral populations were found to correlate with rebound of viral RNA level after prolonged antiretroviral therapies in HIV infected children. Conclusion This rapid and sensitive HIV 1 genotyping method can be used to predict and confirm emergence of HIV 1 drug resistance in HIV infected patients receiving HAART.
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
2000年第4期325-329,共5页
Chinese Journal of Experimental and Clinical Virology
基金
国家自然科学基金资助(3000761748)
关键词
HIV-1
蛋白酶
逆转录酶
序列分析
耐药性
HIV-1
Reverse transcriptase
Protease
Sequence analysis
Drag resistance