Sequence-based genotyping HPV L1 DNA and RNA transcripts in clinical specimens.
We developed a direct sequence-based genotyping method to detect single and multiple HPV L1 DNA and RNA types in genital and dermatological specimens. Our method couples PCR amplification of a highly conserved HPV L1 segment using a broad spectrum-generic primer cocktail mix with automated sequencing of amplified PCR products, followed by GenBank sorting of sequencing data. We genotyped 5 skin and 30 cervical HPV DNA-positive specimens using this method and established its first experimentally derived working cutoff value with the aid of commercial hybridization-based techniques. We suggest that sequence-based genotyping of appropriately amplified DNA and RNA products may serve as a primary HPV detection method in dermatological specimens. It can be applied as an all-purpose genotyping method for rare HPV types not detectable by commercial hybridization-based techniques and for sorting multiple HPV infections by order of prevalence.
Satra
Maria
M
Department of Biology and Genetics, University of Thessalia Medical School, Neo Ktirio, P.O. Box 1400, Larisa 44110, Greece. mariasatra@yahoo.com
Vamvakopoulou
Dimitra N
DN
Sioutopoulou
Despina O
DO
Kollia
Panagoula
P
Kiritsaka
Aspasia
A
Sotiriou
Sotirios
S
Antonakopoulos
Georgios
G
Alexandris
Elias
E
Costantoulakis
Pantelis
P
Vamvakopoulos
Nicholas C
NC
D057205:Automation, Laboratory; D036022:Capsid Proteins / Q000235:genetics*; D002584:Cervix Uteri / Q000821:virology; D003127:Colposcopy; D004279:DNA, Viral / Q000302:isolation & purification*; D030541:Databases, Genetic; D005260:Female; D005838:Genotype; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D030361:Papillomavirus Infections / Q000175:diagnosis / Q000821:virology*; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000302:isolation & purification*; D017422:Sequence Analysis, DNA; D017423:Sequence Analysis, RNA; D012867:Skin / Q000821:virology; D002583:Uterine Cervical Neoplasms / Q000175:diagnosis / Q000821:virology*; D014860:Warts / Q000175:diagnosis / Q000821:virology*; D002578:Uterine Cervical Dysplasia / Q000175:diagnosis / Q000821:virology*
D036022:Capsid Proteins; D004279:DNA, Viral; C114580:HPV L1 protein, Human papillomavirus; D009856:Oncogene Proteins, Viral; D012367:RNA, Viral