尖锐湿疣组织中HPV型别检测及HPV11型L1基因的克隆与测序

目的分析黑龙江地区尖锐湿疣组织感染的HPV型别分布情况,并对HPV11型流行株L1基因进行克隆和测序,检测序列变异情况。方法提取30例尖锐湿疣组织标本中的DNA,用HPVL1区通用引物及特异性HPV6b,11,16,18型引物进行PCR扩增,分析尖锐湿疣组织中感染的HPV型别分布。对其中4株HPV11型阳性标本L1基因,经酶切、连接,分别构建pSP73-HPV11L1重组质粒,并对其L1片段进行测序。结果30例标本中,HPV6b和11型感染数分别为11例,各占36.7%;有2例合并HPV6b/11/16型的混合感染,占总病例数的6.7%;1例HPV16型单独感染,未鉴定出HPV18型感染,其他型别5例。pSP73HPV11L1重组质粒测序后,4株HPV11L1基因序列完全一致,可认为是黑龙江省流行株。其序列与原始株序列相比,仅发现了两处同义点突变。结论黑龙江地区尖锐湿疣组织中HPVDNA检出率为100%,以6b和11型为主,偶见高危型别16型感染。克隆测序表明黑龙江省流行株HPV11型L1基因高度保守。

Analysis of HPV Types and Cloning and Sequencing of HPV 11 L1 Gene in Acuminate Condyloma

Objective To analyze the viral types of human papillomavirus (HPV) in acuminate condyloma biopsy specimens in Heilongjiang province and sequencing of HPV 11 L1 gene.Methods The tissue DNA was extracted from 30 biopsy specimens of acuminate condyloma,respectively.The L1 gene of HPV was amplified with common primers followed by 6b,11,16 and 18 type-specific primers PCR.Four positive L1 fragments of HPV 11 were identified by cloning into plasmid pSP73 and sequencing.Results HPV 6b and HPV 11 were the two most common types,no differences were found between them.Two biopsy specimens were multi-types of HPV 6 b /11/16,amount for 6.7%. In comparison,only one of the 30 biopsy specimens was type 16 (3.3%) and no type 18 was found.And five specimens,16.6% for other HPV types.Sequencing analysis showed the same homology in four clones of HPV type 11 L1 gene,only two synonymous mutations were found compared with the original one in GeneBank.It indicated this local type of HPV 11 represented the epidemic strain in Heilongjiang province.Conclusion The positive rate of HPV infection in all acuminate condyloma biopsy specimens was 100%.The most common types in acuminate condyloma were type 6b and 11,whereas the high-risk type 16 or 18 was very low.High conserved L1 sequence of HPV 11 could represente the epidemic strain in Heilongjiang province.