Antisense RNA directed to the human papillomavirus type 16 E7 mRNA from herpes simplex virus type 1 derived vectors is expressed in CaSki cells and downregulates E7 mRNA

Table 1 Oligonucleotides used for construction of the antisense plasmid and for real-time PCR

Purpose	Name	Sequence (5'-3')
pRB5225	E7ASp-FP	GCTTAGGGTACCATGCATGGAT GATACACCTA
construction	E7ASp-RP	GCTTAGGGTACCCTCTGAGCTGTCATTTAA
Real-time	HBVAS-FP	GAGAAGGGTCGTCCGCAGGAT
PCR	16E7AS-RP	CTCTGAGCTGTCATTTAATTGCTCATA
anti-E7	16E7AS-PRO	(6-FAM)-TGAATATATGTTAGATTTGCAACCAGAGACAACTGATCTCTACT-(TAMRA)
Real-time	16E7-FP	CAGCTCAGAGGAGGAGGATGAA
PCR	16E7-RP	CACACTTGCAACAAAAGGTTACAATATT
HPV-16 E7	16E7-PRO	(6-FAM)-CCAGCTGGACAAGCAGAACCGGAC-(TAMRA)

Abbreviations: AS, antisense; p, plasmid; FP, forward primer, RP, reverse primer; PRO, probe; FAM, 6-carboxyfluorescein; TAMRA, 6-carboxy-tetramethylrhodamine. The underlined base (T) in the primer "E7ASp-FP" is an insertion that introduces a frameshift and two stop codons in the sense orientation of the HPV-16 E7 ORF. The presence of the frameshift mutation in the pRB5225 plasmid was verified by sequencing. The primers and probes for the real-time RT-PCR except for "HBVAS-FP" were designed using Primer Express software (Applied Biosystems).

ISSN: 1743-422X