Medicinal Plant Research 2024, Vol.14, No.6, 345-357 http://hortherbpublisher.com/index.php/mpr 350 Figure 2 CmWD40 regulates the resistance of chrysanthemum plants to Alternaria alternata. (A) The rhythmic expression pattern of CmWD40and the temporal variation in chrysanthemum susceptibility to A. alternata. The line chart represents the rhythmic expression pattern of CmWD40in wild-type (WT) plants under long-day (LD) conditions. Leaves were collected every 6h at the indicated times, and gene expression levels were assessed using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The bar chart represents the mean lesion size on chrysanthemum leaves inoculated with A. alternata at dawn, i.e., at zeitgeber time (ZT)0 and every 6 h for 48 h under LD conditions. The size of the lesions was statistically analyzed at 48 hpi. Data are presented as the mean ± standard error of three biological replicates. (B) Relative expression levels of the CmWD40gene during infection were assessed using qRT-PCR. Data are presented as the mean ± standard error of three biological replicates. Different letters at the top of error bars indicate significant differences (P < 0.05, Tukey’s test). (C) Disease symptoms caused by A. alternata infection in chrysanthemum leaves. Leaves of WT and CmWD40transgenic lines were inoculated withA. alternata, and images were captured at 48 hpi; scale bar=1 cm. (D) Identification of the CmWD40overexpression (OX) transgenic line (OX- CmWD40) at the transcript level using qRT-PCR. Samples were collected at dawn. Different letters at the top of error bars indicate significant differences (P < 0.05, Tukey’s test). (E) Immunoblot analysis of CmWD40in two transgenic lines overexpressing CmWD40. Samples were collected at dawn. Total proteins of transgenic seedlings were extracted and detected with anti-Flag antibodies. Actin was used as an equal loading control. (F) Mean lesion size on WT and OX- CmWD40chrysanthemum leaves inoculated with A. alternataat 48 hpi. Data are presented as the mean ± standard error of three biological replicates. * P ≤ 0.05 compared with control, as calculated by one-way analysis of variance (ANOVA). (G) Expression of CmWD40in CaLCuV-amiR- CmWD40transgenic lines. Samples were collected at dawn. Different letters at the top of error bars indicate significant differences (P < 0.05, Tukey’s test).(H) Disease symptoms caused by A. alternatainfection in chrysanthemum leaves. Leaves of the WT and cabbage leaf-curl geminivirus vector (CaLCuV)-amiR- CmWD40lines were inoculated with A. alternata, and images were captured at 48 hpi; scale bar=1 cm. (I) Mean lesion size on WT and CmWD40silenced chrysanthemum leaves inoculated with A. alternata at 48 hpi. Data are presented as the mean ± standard error of three biological replicates. * P ≤ 0.05 compared with control, as calculated by one-way ANOVA. (J) qRT-PCR analysis for expression of PDF1.2, WRKY33, and MYC2 genes reveals that their transcript levels are upregulated in OX- CmWD40and CaLCuV-amiR- CmWD40transgenic lines, respectively, upon infection with A. alternata. Data are presented as the mean ± standard error of three biological replicates. Different letters at the top of error bars indicate significant differences (P <0.05, Tukey’s test) (Adopted from Zhang et al., 2025)
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