Figure 5.

Expression of DSB repair genes in response to DNA-damaging agents. A, Changes in the transcript abundance of AtPolλ, AtKU80, AtXRCC4, and AtLig4 analyzed by semiquantitative reverse transcription-PCR from 7-d-old wild-type Arabidopsis seedlings without treatment and following exposure to bleomycin (10 μg mL⁻¹) for 8 h (top panels). The − and + symbols indicate the absence and presence of bleomycin. B, Expression of core NHEJ-related DSB repair genes AtKU80, AtXRCC4, and AtLig4 in 7-d-old wild-type Arabidopsis seedlings after treatments with DNA-damaging agents. Lane 1 in each panel indicates mRNA levels of the respective genes in untreated control seedlings, while lanes 2 to 4 indicate expression of the genes after exposure of seedlings to 200 mm NaCl, 80 µL L⁻¹ MMS, and 5 μg mL⁻¹ MMC, respectively, for 8 h. C and D, Changes in mRNA levels of NHEJ genes in 7-d-old wild-type seedlings in various recovery time periods after exposure to 200 mm NaCl (C) and 5 μg mL⁻¹ MMC (D) for 8 h. The 0-h time point served as the control (lane 1). The AtPolλ loss-of-function mutation reduces the expression of core DSB repair genes. E, Expression of NHEJ genes in 7-d-old wild-type and atpolλ mutant seedlings. F and G, Changes in mRNA levels of NHEJ genes in 7-d-old atpolλ-1 mutant seedlings during various recovery time periods after treatment with 200 mm NaCl (F) and 5 μg mL⁻¹ MMC (G) for 8 h. H and I, Expression of NHEJ genes in 7-d-old atpolλ-2 mutant seedlings during posttreatment recovery periods after exposure to 200 mm NaCl (H) and 5 μg mL⁻¹ MMC (I) for 8 h. The 0-h time point served as the control (lane 1). Expression of β-tubulin is shown as a control (bottom panels). Quantification of the changes in transcript levels is shown in Supplemental Figure S12 for B to D and in Supplemental Figure S13 for E to I. Specificity of the amplification of transcripts was confirmed by direct sequencing of the gel-purified PCR products.