Human papillomavirus detection and typing using a nested-PCR-RFLP assay
Background: It is clinically important to detect and type human papillomavirus (HPV) in a sensitive and specific manner. Objectives: Development of a nested-polymerase chain reaction-restriction fragment length polymorphism (nested-PCR-RFLP) assay to detect and type HPV based on the analysis of L1 g...
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doaj-2752437273c04c178b4f18315b277ba82020-11-25T02:58:33ZengElsevierBrazilian Journal of Infectious Diseases1413-86702011-09-01155467472Human papillomavirus detection and typing using a nested-PCR-RFLP assayJanaina Coser, MSc0Thaís da Rocha Boeira, MSc1André Salvador Kazantzi Fonseca, MSc2Nilo Ikuta, PhD3Vagner Ricardo Lunge, PhD4Professor, Postgraduate Program in Genetics and Molecular Diagnosis, Universidade Luterana do Brasil (ULBRA), Canoas, RS, Brazil, and Universidade de Cruz Alta (UNICRUZ), Cruz Alta, RS, BrazilTechnical Manager, Simbios Biotecnologia, Canoas, RS, BrazilResearcher, Simbios Biotecnologia, Canoas, RS, BrazilProfessors, Postgraduate Program in Genetics and Molecular Diagnosis, ULBRA, Canoas, RS, Brazil; Simbios Biotecnologia, Canoas, RS, BrazilProfessors, Postgraduate Program in Genetics and Molecular Diagnosis, ULBRA, Canoas, RS, Brazil; Simbios Biotecnologia, Canoas, RS, Brazil; Correspondence to: ULBRA Laboratório de Diagnóstico Molecular Avenida Farroupilha, 8001 Prédio 22 - sala 312 Bairro São José 92425-900, Canoas, RS, Brazil.Background: It is clinically important to detect and type human papillomavirus (HPV) in a sensitive and specific manner. Objectives: Development of a nested-polymerase chain reaction-restriction fragment length polymorphism (nested-PCR-RFLP) assay to detect and type HPV based on the analysis of L1 gene. Methods: Analysis of published DNA sequence of mucosal HPV types to select sequences of new primers. Design of an original nested-PCR assay using the new primers pair selected and classical MY09/11 primers. HPV detection and typing in cervical samples using the nested-PCR-RFLP assay. Results: The nested-PCR-RFLP assay detected and typed HPV in cervical samples. Of the total of 128 clinical samples submitted to simple PCR and nested-PCR for detection of HPV, 37 (28.9%) were positive for the virus by both methods and 25 samples were positive only by nested-PCR (67.5% increase in detection rate compared with single PCR). All HPV positive samples were effectively typed by RFLP assay. Conclusion: The method of nested-PCR proved to be an effective diagnostic tool for HPV detection and typing. Keywords: DNA probes, HPV, polymerase chain reaction, polymorphism, restriction, fragment lengthhttp://www.sciencedirect.com/science/article/pii/S141386701170229X |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Janaina Coser, MSc Thaís da Rocha Boeira, MSc André Salvador Kazantzi Fonseca, MSc Nilo Ikuta, PhD Vagner Ricardo Lunge, PhD |
spellingShingle |
Janaina Coser, MSc Thaís da Rocha Boeira, MSc André Salvador Kazantzi Fonseca, MSc Nilo Ikuta, PhD Vagner Ricardo Lunge, PhD Human papillomavirus detection and typing using a nested-PCR-RFLP assay Brazilian Journal of Infectious Diseases |
author_facet |
Janaina Coser, MSc Thaís da Rocha Boeira, MSc André Salvador Kazantzi Fonseca, MSc Nilo Ikuta, PhD Vagner Ricardo Lunge, PhD |
author_sort |
Janaina Coser, MSc |
title |
Human papillomavirus detection and typing using a nested-PCR-RFLP assay |
title_short |
Human papillomavirus detection and typing using a nested-PCR-RFLP assay |
title_full |
Human papillomavirus detection and typing using a nested-PCR-RFLP assay |
title_fullStr |
Human papillomavirus detection and typing using a nested-PCR-RFLP assay |
title_full_unstemmed |
Human papillomavirus detection and typing using a nested-PCR-RFLP assay |
title_sort |
human papillomavirus detection and typing using a nested-pcr-rflp assay |
publisher |
Elsevier |
series |
Brazilian Journal of Infectious Diseases |
issn |
1413-8670 |
publishDate |
2011-09-01 |
description |
Background: It is clinically important to detect and type human papillomavirus (HPV) in a sensitive and specific manner. Objectives: Development of a nested-polymerase chain reaction-restriction fragment length polymorphism (nested-PCR-RFLP) assay to detect and type HPV based on the analysis of L1 gene. Methods: Analysis of published DNA sequence of mucosal HPV types to select sequences of new primers. Design of an original nested-PCR assay using the new primers pair selected and classical MY09/11 primers. HPV detection and typing in cervical samples using the nested-PCR-RFLP assay. Results: The nested-PCR-RFLP assay detected and typed HPV in cervical samples. Of the total of 128 clinical samples submitted to simple PCR and nested-PCR for detection of HPV, 37 (28.9%) were positive for the virus by both methods and 25 samples were positive only by nested-PCR (67.5% increase in detection rate compared with single PCR). All HPV positive samples were effectively typed by RFLP assay. Conclusion: The method of nested-PCR proved to be an effective diagnostic tool for HPV detection and typing. Keywords: DNA probes, HPV, polymerase chain reaction, polymorphism, restriction, fragment length |
url |
http://www.sciencedirect.com/science/article/pii/S141386701170229X |
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