| 张佳瑞 巩丽 朱少君 韩秀娟 姚丽 王姝妹 李艳红 张伟.实时定量RT-PCR对乙型肝炎病毒全长RNA(fRNA)的定量检测[J].现代生物医学进展英文版,2015,15(1):29-32. |
| 实时定量RT-PCR对乙型肝炎病毒全长RNA(fRNA)的定量检测 |
| Validation of a Simple Real-time RT-PCR for Detection and Quantizationof HBV fRNA |
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| DOI： |
| 中文关键词: HBV RNA 实时荧光定量 fRNA Hayashi's定量法 |
| 英文关键词: HBV RNA RT-PCR fRNA Hayashi's quantification method |
| 基金项目:国家自然科学基金项目（30672013）；国家自然科学基金项目（81372226）；陕西省科技统筹创新工程（2011KTCL03-11） |
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| 中文摘要: |
| 目的：建立一种简便的定量检测慢性乙型肝炎患者血清中终止于聚腺苷酸化位点的乙型肝炎病毒全长RNA(fRNA)的方法。
方法：选取53 例未治疗的乙型肝炎患者及22 例HBsAg阴性的健康者为研究对象，使用锚定oligo-dT 的引物对其血清中fRNA
进行实时定量反转录PCR 检测，统计分析其与HBV DNA、HBcrAg 和HBeAg 的相关性。结果：对fRNA 进行实时荧光定量
RT-PCR 检测的下线为2.3 log copies/ml，标准曲线的相关系数为0.99(P<0.0001)。53 例乙型肝炎患者中，29 例(54.7%)可以检测到
fRNA，22 例正常对照中没有检测到fRNA。27 例HBeAg 阳性和/ 或高水平HBV DNA的患者全部检测到fRNA，26例HBeAg阴
性并且低水平HBV DNA 的乙型肝炎患者中有2 例(7.7%) 检测到fRNA (P<0.0001)。HBeAg 阳性患者血清中fRNA水平高于
HBeAg阴性患者(5.0± 0.3 vs. 2.9± 0.4 log copies/ml, P<0.001)。fRNA与HBV DNA/HBcrAg 具有显著相关性(r=0.905、0.881，P<0.
0001)。Hayashi's 定量分析法I显示fRNA与HBV DNA 相关性强于其与HBcrAg 的相关性。结论：与HBV DNA 和HBeAg 一
样，fRNA 可作为常规检测判断HBV的复制水平并指导用药。 |
| 英文摘要: |
| Objective:To establish a simple assay for the detection and quantization of full-length RNA (fRNA) terminating at
polyadenilation site in sera of chronic hepatitis B (CHB) patients.Methods:fRNA were assayed via TaqMan real-time RT/PCR using anchored
oligo-dT primers in sera of 53 treatment-naive CHB patients and 22 HBsAg-negative healthy controls. Results were analyzed by
comparation of HBV DNA with HBcrAg and HBeAg.Results:The fRNA assay had a lower limit of detection and quantization at 2.3 log
copies/ml, and a correlation coefficient of 0.99 (P<0.0001). fRNA was detected in 29 of 53 (54.7%) of the CHB patients as compared to
non of 22 controls (Specificity). fRNA was detected in all 27 HBeAg-positive and/or high HBV DNA levels CHB patients as compared to
2 of 26 (7.7%) HBeAg-negative and low HBV DNA levels CHB patients (P<0.0001). fRNA levels were higher in HBeAg-positive than
in HBeAg-negative samples (5.0± 0.3 vs. 2.9± 0.4 log copies/ml, P<0.001). Significant correlation was found between fRNA and HBV
DNA/ HBcrAg (r=0.905 and 0.881, respectively, P<0.0001). The effective items on fRNA levels, in descending order, were: HBV DNA,
HBcrAg by means of Hayashi's quantification method type I (Multiple correlation efficient=0.939).Conclusion:The simple real-time
RT/PCR for detection and quantization of fRNA was suitable for routine clinical test in assessing HBV replication status the same as
HBV DNA and HBeAg in CHB patients. |
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