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. 2001 Oct 30;98(23):13437–13442. doi: 10.1073/pnas.241276898

Figure 5.

Figure 5

Comparison of ipt and iaaM transcript abundance in wild-type and transgenic tomato by using competitive reverse transcriptase–PCR. Wild-type plants and plants from the crown gall-resistant transgenic line 01/6 were stem inoculated with A. tumefaciens 20W-5A, and RNA was extracted from inoculated stem tissue 3, 10, and 20 dpi. First-strand cDNA synthesis was primed with a polyT primer, and cDNA was subsequently PCR amplified in the presence of a known concentration of serially diluted competitive standard. (A) Agarose gel electrophoresis of 20 dpi iaaM competitive PCR reactions. Note the presence of an 815-bp internal standard amplification product (St) and a 1,035-bp iaaM cDNA amplification product (M). (B) Agarose gel electrophoresis of 20 dpi ipt competitive PCR reactions. Note the presence of an 815-bp internal standard amplification product (St) and a 609-bp ipt cDNA amplification product (T). Lane markers apply to both panels. Lane a, molecular weight standard; lanes b–d, PCR amplification of wild-type cDNA in the presence of 0.002 pg (lane b), 0.01 pg (lane c), and 0.05 pg (lane d) of iaaM internal standard (A) or 0.005 pg (lane b), 0.05 pg (lane c), and 0.5 pg (lane d) of ipt internal standard (B). Lanes e–g, PCR amplification of cDNA from the transgenic line 01/6 in the presence of 0.002 pg (lane e), 0.01 pg (lane f), and 0.05 pg (lane g) of iaaM internal standard (A) or 0.005 pg (lane e), 0.05 pg (lane f), and 0.5 pg (lane g) of ipt internal standard (B). Lane H, PCR amplification of cDNA from a noninoculated control plant. (C) Comparison of oncogene mRNA abundance in wild-type and 01/6 plants at 3, 10, and 20 dpi. *, relative mRNA suppression refers to the abundance of 01/6 cDNA oncogene PCR amplification product as compared with the wild-type cDNA oncogene product (1 − [conc. of 01/6 product/conc. of wild-type product]). †, examples in which the 01/6 cDNA oncogene product was below the lower limit of detection are noted; the concentration of these products was recorded as less than the lowest concentration of internal standard.