Expression of a LMP2 transcript containing exons 2 to 6 in EBV positive NK- and T-cell tumor lines. (A) End point PCR products (35 cycles) visualized on a 1% agarose gel. Amplification of cDNA using a forward primer located in exon 1 and reverse primer within exon 3 (F1 and R3; see supplemental Table 1) yields a product of expected size in the LCL but no product in the NK- and T-cell lines. PCR performed with a forward primer in exon 2 and a reverse primer located within exon 6 (F2 and R6) produces a product of the anticipated size in all 4 NK- and T-cell lines. The EBV negative NK leukemia cell line, NKL, and a no reverse transcriptase (NO RT) sample from a LCL served as negative controls. (B) Quantitative RT-PCR assays, using primers and probes designed to amplify LMP2 transcripts containing exons 2 and 6, respectively (avoiding exon-exon junctions in the event of uncharacterized splice variations). Data are expressed using cycle threshold (Ct) values, where Ct₀ represents the assay threshold of detection. LMP2 transcript Ct values are normalized to cellular glyceraldehyde 3-phosphate dehydrogenase (GAPDH) Ct values. The left-hand y-axis represents Δ(Ct₀-Ct), while the right-hand axis shows the relative transcript levels.