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. 2019 Feb 11;116(9):3494–3501. doi: 10.1073/pnas.1819769116

Fig. 2.

Fig. 2.

HAN1 is a negative regulator of chilling tolerance. (A) The DNA construct used for estrogen-inducible overexpression of HAN1 in rice. VEX is the gene coding a chimeric transcriptional activator that binds the pER8 promoter, which consists of eight copies of the LexA operator fused upstream of the −46 35S minimal promoter, to activate transcription of HAN1 in the presence of exogenous estrogen. (B and C) Performance of pER8::HAN102428 transgenic (No. 1–3) and wild-type (WT) lines before (B) and after chilling treatment (C). (D) Expression levels of HAN1 in pER8::HAN102428 transgenic lines and wild type. (E) Survival rate (%) of pER8::HAN102428 transgenic lines and wild type after chilling stress treatment. The DNA construct designed for knockout mutation of HAN1 using the CRISPR/Cas9 system. (G–H) Performance of knockout han1 mutant (han1-1 and han1-2) and wild-type (NP) lines before (G) and after chilling treatment (H). (I) Partial DNA sequences from the HAN1 knockout mutants (han1-1 and han1-2) show one base (A or T) insertion in CDS region of HAN1 for the two alleles, resulting in frameshift mutations. (J) Survival rate (%) of HAN1 knockout mutants and wild-type control after chilling treatment. Significant differences were determined by one-way ANOVA (*P < 0.05, or **P < 0.01, n = 3) in D, E, and G.