Design of qPCR primers for the early region of Human Papillomavirus oncogenic types 16 and 18

Primer pairs for the HR-HPV16 and 18 were designed to target the open reading frames (ORF) E6, E7, E1 and E2 in real-time PCR assays. This data is provided in the folders: Figures, Tables and HPV Alignments

1. HPV alignments folder contain the following files:
For gene-specific qPCR primer design, poorly conserved nucleotide regions were identified from HPV alignments of full-length genomes of types HPV16, 31, 35 and HPV 18, 45 and 59 in FASTA format. The HPV clusters are available: “HPV16_31_35_alignment.fa”; “HPV18_45_59_alignment.fa”.

2. A list of qPCR primers proposed is presented in Table 1. A graphic representation of amplification strategy is shown in Figure 1. Briefly, primers that produce overlapping amplicons less than 600 bp, that passed the secondary structure tests, designed with annealing temperatures above 58°C, were selected for experimental evaluation. Calibration curves were generated using SYBR Green chemistry and serial dilutions of a DNA standard as template for each genotype evaluated. For ORF HPV18 E7, the amplification plots, including the melt curve analysis are shown in Figure 2. Primer pairs producing multiple products (Figure 3), are not recommended. Complete information for primer pair used in real-time PCR assays are described in Table 2. All this information is presented in Figures and Tables folders.