Extended Data Fig. 6: Sequence of SMPED, overexpression/knockdown of SMPEDs in Arabidopsis thaliana and validation of promoters.
a, Sequence of AmSMPED1 and its two homologous proteins in Arabidopsis thaliana (At5g67020 and At3g50340). b, Confirmation of overexpression of AmSMPED1 (35S:AmSMPED1), AtSMPED1 (35S:AtSMPED1), and AtSMPED2 (35S:AtSMPED2), as well as simultaneous knockdown of AtSMPED1 and AtSMPED2 (ami-AtSMPED1/2) in A. thaliana. Full-length amplification of AmSMPED1 was then performed, and the results showed that, under identical starting RNA concentrations, the amplified bands from overexpression lines were notably brighter compared to those from wild-type and positive control plants, indicating successful overexpression of AmSMPED1 in the transgenic lines. Overexpression of AmSMPED1 (35S:AmSMPED1) was confirmed by RT-PCR. Total RNA from wild-type plants, positive controls, and transgenic lines was quantified and normalized to the same initial concentration. The data are presented as the mean ± SEM and the whiskers extend to minima and maxima. Biological replicates (n) of each treatment are indicated. The blank space on the bar indicates contracted bar due to large values. The asterisks denote significant differences between samples using two-sided Mann–Whitney U-test. ***, P < 0.0001. c, Validation of promoters in the DDR. The upper diagram is the location of promoters in the DDR. “ATG” is the initiation codon of AmSMPED1. The lower is a diagram for validation. LUC is luciferase and REN is Renilla luciferase. The results of validation are indicated in Fig. 3b.
