Figure 1
Generation of transgenic mice. (A) Bacterial artificial chromosome encoding wild-type mouse genomic Tmc1 (Tmc1 BAC) was used to construct Tg[Tmc1∆Ex8_9], Tg[PTmc1::Tmc2] or Tg[PTmc1::Tmc1]. To construct Tg[Tmc1∆Ex8_9], we deleted 2775 bp including exons 8 and 9. This deleted region is identical to that deleted in Tmc1∆/∆ mice14. To construct Tg[PTmc1::Tmc2] or Tg[PTmc1::Tmc1], we modified Tg[Tmc1∆Ex8_9] to replace the translation initiation codon of Tmc1 with the Tmc1 or Tmc2 cDNA conjugated with the SV40 polyadenylation signal, respectively. (B) Relative total Tmc2 mRNA levels (mean ± SD) of Tg[PTmc1::Tmc2]; Tmc1∆/∆ cochleae (black line) and of wild-type cochleae (Tmc1+/+, gray line). In Tg[PTmc1::Tmc2]; Tmc1∆/∆ cochleae, transgenic Tmc2 mRNA (dashed line) was identified after E17.5 and remained between 8- and 16-fold levels at P25, the last time point we examined. Endogenous Tmc2 mRNA (dotted line) was almost identical to those of wild-type cochleae. The number above each data point and bar indicates the number of mice examined. Relative RNA level was calculated by normalizing the level of each mRNA for each time point to the Actb level at that time point and then to the normalized Tmc2 mRNA level in wild-type cochlea measured at P0.
