RT-PCR analysis for the internal priming (IP) candidates in fetal liver, colon and lung total RNA. RT-PCR was conducted in DNA-free RNA previously treated with DNAse (lanes 1 and 2) and in untreated RNA, which was, therefore, contaminated with genomic DNA (gDNA; lanes 3 and 4) for each candidate in the corresponding tissue. As a control, RT-PCR was conducted in the presence (lanes 1 and 3) and absence (lanes 2 and 4) of reverse transcriptase. gDNA was used as a positive control of the PCR reaction (lane 5) and no template as a negative control (lane 6). For fetal liver, in 3 IP candidates (5, 8 and 11) the PCR products (152 bp, 153 bp and 160 bp, respectively) were observed in the treated RNA when RT was added (lane 1) or in untreated RNA independent of the RT (lanes 3 and 4). For colon, in 1 IP candidate (9) the PCR product (158 bp) was observed in the treated RNA when RT was added (lane 1) or in untreated RNA independent of the RT (lanes 3 and 4). For the remaining IP candidates (1, 2, 4, 6, 7, 10 and 12), the PCR products (214 bp, 229 bp, 207 bp, 156 bp, 227 bp, 205 bp and 234 bp, respectively) were observed only in untreated RNA independent of the RT (lanes 3 and 4). The PCR products were analyzed on 8% polyacrylamide gels with silver staining. A 100 bp ladder (M) was used as molecular weight marker. In each gel the lower fragment in lane M correspond to 100 bp.
Galante et al. Genome Biology 2007 8:R40 doi:10.1186/gb-2007-8-3-r40