丙型肝炎病毒不同基因区的聚合酶链反应扩增  

Detection of hepatitis C virus by polymerase chain reaction with primers deduced from different gene region

作　　者：魏来[1] 陶其敏[1] 杜绍财[1] 王豪[1] 孙焱[1] 

机构地区：[1]北京医科大学人民医院肝病研究所

出　　处：《中华医学检验杂志》1997年第6期346-348,共3页

基　　金：国家八五攻关及CMB基金

摘　　要：目的探讨提高聚合酶链反应（ＰＣＲ）对丙型肝炎病毒（ＨＣＶ）病毒血症的检出率和对变异较大区域的扩增效率的方法。方法分别以ＨＣＶ基因组５＇端非编码区（５＇ＵＴＲ）、非结构基因４区（ＮＳ４）及ＮＳ５ｂ区的引物检测ＨＣＶ，并比较双退火温度与单一退火温度对ＨＣＶ基因扩增效率的影响。结果ＨＣＶ不同基因区（５′ＵＴＲ、ＮＳ４、ＮＳ５ｂ）引物的ＰＣＲ扩增阳性率分别为９２％、６２％、５９％。以双退火温度扩增ＮＳ４区与ＮＳ５ｂ区基因，扩增阳性率分别提高到７７％与７９％。将血清按原血清、１０－１～１０－１０系列稀释后检测，发现２份血清单退火温度ＰＣＲ的检出水平分别为１０－５与１０－７，双退火温度ＰＣＲ检出水平则分别提高至１０－８与１０－９稀释血清。结论５′ＵＴＲ引物对ＨＣＶＲＮＡ的检出率明显高于ＮＳ４区与ＮＳ５ｂ区引物。Objective To investigate the effect of variability of hepatitis C virus (HCV) on detection of HCV with polymerase chain reaction (PCR), and establish a method to improve the sensitivity and effectiveness of detection. Methods Detection of HCV was performed with three pairs primers deduced from 5′untranslated region (5′UTR), nonstructural gene 4 (NS4), and nonstructural gene 5 (NS5) of HCV. Positive rates were compared. In addition, the positive rate between one anneling temperature PCR and two annealing temperature PCR was compared as well. Results The positive rates of detection with primers deduced from 5′UTR, NS4 and NS5 were 92%, 62% and 59% respectively. With two annealing temperature PCR, the positive rates were increased to 77% and 79% respectively by primers of NS4 and NS5. In detection by one annealing temperature, detectable levels of 10 5 and 10 7 diluted sera were obtained from sample 1 and sample 2. However, detectable levels were 10 8 and 10 9 diluted sera with two annealing temperature PCR. Conclusion Detection of HCV is more sensitive with primers of 5′UTR than with NS4 or NS5 primer. Two annealing temperature PCR is a candidate for increasing positive rate of variable gene of HCV.

关 键 词：丙型肝炎病毒 基因 病毒 聚合酶链反应 

分 类 号：R512.630.4[医药卫生—内科学] R446.9[医药卫生—临床医学]