Journal: Journal of Experimental Botany

Article Title: Arabidopsis SKP1-like protein13 (ASK13) positively regulates seed germination and seedling growth under abiotic stress

doi: 10.1093/jxb/ery191

Figure Lengend Snippet: Quantitative RT-PCR analysis of ASK13 ( At3g60010 ) in (A) leaves, roots, stem, and flowers of 6-week-old mature plants, siliques with seeds at 14–16 d after pollination (DAP), and seeds after-ripened for 8 weeks; (B) during seed development; (C) during seed germination; (D) in 7-d-old seedlings exposed to various forms of abiotic stress; and (E) in 7-d-old seedlings treated with various phytohormones (Con, control; SA, salicylic acid; BAP, 6-benzylaminopurine; IAA, indole-3-acetic acid; ABA, abscisic acid; ACC, 1-aminocyclopropane-1-carboxylic acid; JA, methyl jasmonate; GA, gibberellic acid). Total RNA from each sample was reverse-transcribed and subjected to real-time PCR analysis. The relative expression value of each gene was normalized using 18S ribosomal small subunit RNA as the endogenous control, and calculated using the 2 –ΔΔ C T method . Data are means (±SD) from triplicate analyses of three biological replicates. Different letters indicate significant differences according to Duncan’s Multiple Range Test ( P <0.01). (F) Subcellular localization of ASK13 as determined using green fluorescent protein (GFP). Images show localization of GFP and ASK13-GFP in roots of 5-d-old transgenic Arabidopsis seedlings. Cells were stained with propidium iodide (PI), and the emission signal was detected between 580–680 nm. Bright-field and merged images are also shown.

Article Snippet: The pGBKT7BD- ASK13 transformed cells were then mated with a normalized Arabidopsis Y2H cDNA library (purchased from Clonetech) and screening was carried out according to the manufacturer’s guidelines.

Techniques: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Expressing, Transgenic Assay, Staining

Journal: Journal of Experimental Botany

Article Title: Arabidopsis SKP1-like protein13 (ASK13) positively regulates seed germination and seedling growth under abiotic stress

doi: 10.1093/jxb/ery191

Figure Lengend Snippet: List of ASK13-interacting proteins and their functions

Article Snippet: The pGBKT7BD- ASK13 transformed cells were then mated with a normalized Arabidopsis Y2H cDNA library (purchased from Clonetech) and screening was carried out according to the manufacturer’s guidelines.

Techniques: Binding Assay, Expressing

Journal: Journal of Experimental Botany

Article Title: Arabidopsis SKP1-like protein13 (ASK13) positively regulates seed germination and seedling growth under abiotic stress

doi: 10.1093/jxb/ery191

Figure Lengend Snippet: ASK13 -overexpression results improved Arabidopsis seed germination, vigor, and seedling growth under abiotic stress conditions. Results from three representative independent transformed lines are shown ( ASK13 -OE 1–3). Seeds that had been after-ripened for 8 weeks were used for these experiments, and all data are means (±SD) of four replicates with 50 seeds each. (A) Germination percentage of wild-type ( WT ), empty vector ( VC ), and ASK13 -transformed seeds. (B) Comparison of germination percentage among seeds under various stress conditions. (C) Germination percentage of seeds without (0 d) or with controlled deterioration treatment (CDT) applied for 4 d. Germination was scored after 7 d of imbibition following the CDT. (D) Viability of seeds with or without CDT, analysed using tetrazolium staining: dark red indicates viable seeds. The images of each genotype were taken separately. Quantitative analysis of (E) H 2 O 2 and (F) malondialdehyde (MDA) content in the seeds with or without CDT. In the graphs, different letters indicate significant differences among means according to Duncan’s Multiple Range Test ( P <0.01). (G) Phenotypes of seedlings under various forms of abiotic stress. Stress treatments were applied to 7-d-old seedlings: cold stress, 4 °C; osmotic stress, PEG, –0.5MPa; heat stress, 37 °C; salinity stress, NaCl, 150 mM; and oxidative stress, 2 μM paraquat.

Article Snippet: The pGBKT7BD- ASK13 transformed cells were then mated with a normalized Arabidopsis Y2H cDNA library (purchased from Clonetech) and screening was carried out according to the manufacturer’s guidelines.

Techniques: Over Expression, Transformation Assay, Plasmid Preparation, Staining

Journal: Journal of Experimental Botany

Article Title: Arabidopsis SKP1-like protein13 (ASK13) positively regulates seed germination and seedling growth under abiotic stress

doi: 10.1093/jxb/ery191

Figure Lengend Snippet: The ask13 knockdown mutant of Arabidopsis shows reduced seed germination, vigor, and seedling growth under abiotic stress conditions. Seeds that had been after-ripened for 8 weeks were used for these experiments, and all data are means (±SD) of four replicates with 50 seeds each. (A) Germination percentage of wild-type (WT) and ask13 knockdown seeds. (B) Germination percentage under various stress conditions. (C) Germination percentage without (0 d) or with controlled deterioration treatment (CDT) applied for 4 d. Germination was scored after 7 d of imbibition following the CDT. (D) Viability of seeds with or without CDT, analysed using tetrazolium staining: dark red indicates seeds are viable. The images of each genotype were taken separately. Quantitative analysis of (E) H 2 O 2 and (F) malondialdehyde (MDA) content in seeds with or without CDT. In the graphs, different letters indicate significant differences among means according to Duncan’s Multiple Range Test ( P <0.01). (G) Phenotypes of seedlings under various forms of abiotic stress. Stress treatments were applied to 7-day-old seedlings: cold stress, 4 °C; osmotic stress, PEG, –0.5MPa; heat stress, 37 °C; salinity stress, NaCl, 150mM; and oxidative stress, 2 μM paraquat.

Article Snippet: The pGBKT7BD- ASK13 transformed cells were then mated with a normalized Arabidopsis Y2H cDNA library (purchased from Clonetech) and screening was carried out according to the manufacturer’s guidelines.

Techniques: Mutagenesis, Staining