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首页> 外文期刊>Journal of Clinical Microbiology >Comparison of Variant-Specific Hybridization and Single-Strand Conformational Polymorphism Methods for Detection of Mixed Human Papillomavirus Type 16 Variant Infections
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Comparison of Variant-Specific Hybridization and Single-Strand Conformational Polymorphism Methods for Detection of Mixed Human Papillomavirus Type 16 Variant Infections

机译：变异特异性杂交和单链构象多态性检测混合型人乳头瘤病毒16型变异感染的比较

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PCR-based variant-specific hybridization (VSH) and single-strand conformational polymorphism (SSCP) analyses were compared for their capacities to detect mixed human papillomavirus type 16 (HPV-16) variant infections within clinical specimens. The SSCP assay used in this comparison targets a 682-bp fragment that spans nucleotides 7445 to 222 within the HPV-16 genome. This fragment includes portions of the HPV-16 long control region and the E6 open reading frame and identifies three categories of SSCP patterns: those identical to the patterns of prototype HPV-16 (P), those identical to the patterns of Caski-derived HPV-16 (C), or those that are different from the P and C HPV-16 patterns and that are therefore classified as belonging to novel (N) HPV-16 variants. VSH targets the entire HPV-16 E6-coding region (nucleotides 56 to 640) and distinguishes previously described variant nucleotides at positions 109, 131, 132, 143, 145, 178, 286, 289, 350, 403, and 532. Clinical samples used in VSH and SSCP analyses were subjected to multiple independent amplification reactions. The resultant amplicons were cloned, and 14 to 78 clones per clinical specimen were evaluated by VSH. VSH detected an HPV-16 variant that represented at least 20% of the amplified HPV-16 variant population. In contrast, SSCP analysis detected HPV-16 variants that represented 36% of the amplified HPV-16 population. Comparison studies were conducted with mixed HPV-16 variant laboratory constructs. Again, VSH had a higher sensitivity than SSCP analysis in detecting mixed HPV-16 variant infections in these constructed amplicon targets. Accurate detection of HPV-16 variants may enhance our understanding of the natural history of HPV-16 infections.
机译：比较了基于PCR的变异特异性杂交（VSH）和单链构象多态性（SSCP）分析在临床标本中检测混合型16人乳头瘤病毒（HPV-16）变异感染的能力。在此比较中使用的SSCP分析的目标是一个682bp的片段，该片段跨越HPV-16基因组内的7445至222位核苷酸。该片段包括HPV-16长控制区和E6开放阅读框的部分，并确定了三类SSCP模式：与原型HPV-16（P）模式相同的那些，与从Caski衍生的HPV模式相同的那些-16（C）或不同于P和C HPV-16模式的那些，因此被归类为属于新型（N）HPV-16变体。 VSH靶向整个HPV-16 E6编码区（核苷酸56至640），并在位置109、131、132、143、145、178、286、289、350、403和532处区分先前描述的变异核苷酸。在VSH和SSCP分析中使用的cDNA进行了多个独立的扩增反应。克隆得到的扩增子，并通过VSH评估每个临床样品14至78个克隆。 VSH检测到HPV-16变异体，该变异体至少占扩增的HPV-16变异体群体的20％。相比之下，SSCP分析检测到HPV-16变异体，占变异的HPV-16群体的36％。使用混合的HPV-16变异实验室构建物进行了比较研究。同样，VSH在检测这些构建的扩增子靶标中混合的HPV-16变异感染方面比SSCP分析具有更高的灵敏度。准确检测HPV-16变异可能会增强我们对HPV-16感染自然史的理解。

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