For MTT assays, cells (2,000 ~ 5,000 cells/well) were subcultured into 96-well plates according to their growth properties. Cell proliferation was assayed at 72 hr after treatment of CHIR-99021 by adding 20 μl of 5 mg/ml 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) solution per 100 μl of growth medium. After incubating for 3-4 h at 37°C, the media were removed and 150 µl/well of MTT solvent (either absolute DMSO or isopropanol containing 4 mM HCl and % Nonidet-40) was added to dissolve the formazan. The absorbance of each well was measured by ELx808 (BioTek, Winooski, VT) or Wallac Victor2 (Perkin-Elmer Life Sciences, Boston, MA) Microplate Reader. Viable cells are presented as percent of control, vehicle-treated cells.
Seed treatment is a useful practice for healthy plant growth. Seed priming through controlled hydration and dehydration enhances early germination of mustard seed in less time, even in compacted soil [ 8 ]. The soaking of mustard seeds in % aqueous pyridoxine hydrochloride solution for 4 hours improved germination. The combination of pyridoxine + N 60 P 20 + N 15 P 5 (top dressing) accelerated the crop performance by enhancing seed yield and oil yield by and %, respectively, over the control [ 9 ]. The differential response of varieties for imbibition gives advantage to some of them to germinate early as compared to others. At Hisar, maximum rate of imbibition was reported in “NRCDR-2” (%) and minimum in “NRCDR-509” (%). Such drastic difference in rate of imbibition is important for identification of suitable varieties under abiotic stress conditions namely drought, frost, and temperature abnormalities.
Seed germination performance is a major determinant of crop yield. Deterioration of seed quality with age is associated with accumulation of DNA damage .  In dry, aging rye seeds, DNA damages occur with loss of viability of embryos.  Dry seeds of Vicia faba accumulate DNA damage with time in storage, and undergo DNA repair upon germination.  In Arabidopsis , a DNA ligase is employed in repair of DNA single- and double-strand breaks during seed germination and this ligase is an important determinant of seed longevity.  In eukaryotes , the cellular repair response to DNA damage is orchestrated, in part, by the DNA damage checkpoint kinase ATM . ATM has a major role in controlling germination of aged seeds by integrating progression through germination with the repair response to DNA damages accumulated during the dry quiescent state.