Nonquantitative, sequence-independent RT-PCR (SIP-RT-PCR) of single daughter cells induced to unilineage erythroid differentiation. A representative experiment is shown. (Upper panel) The mRNA isolated from a single round III cell was processed for SIP-RT-PCR or SIP-RT-PCR was performed on whole cell lysate (w.c.l.; upper panel right lane). PCR products were then hybridized with32P-labeled probes (upper panel). Autoradiographs are shown. (Lower panel) Autoradiographs of corresponding sequence-specific PCR products from SIP-RT-PCR products shown in the upper panel. SIP-RT-PCR product (2.5 μL) was reamplified by PCR using sequence-specific pairs of primers (SSP-RT-PCR) recognizing the respective target gene. Note that the whole cell lysate (w.c.l.) SIP-RT-PCR results in coamplification of genomic DNA (outermost right lane) by using intron-spanning primers for β-microglobulin. No PCR products corresponding to genomic β2-microglobulin DNA sequences were detected when SIP-RT-PCR product from mRNA was reamplified by the same β2-microglobulin specific primers (lower panel, second lane from the right).
