Three homozygous T3 lines (5T, 20T and 8T) expressing
PSARK::IPT alongside with nulls and WT plants were germinated
on moist germination paper for 6 days (30 C in dark). Seedlings
were then transplanted into 4.5-L pots, filled with soil (capay
series, harvested in California rice field,
3832¢23.93¢¢N,12148¢30.81¢¢W, shredded and steamed for
1.5 h to eradicate soil pathogens), with two plants per pot, and
placed in water tubes. Greenhouse conditions were kept 12 h,
30 C (day) ⁄ 12 h, 20 C (night). Plants were fertilized with a
solution 50% N:P:K (20:10:20) and 50% ammonium sulphate
(total of 0.5 g nitrogen) every 10 days until panicle initiation.
Drought treatments were applied at two developmental stages:
pre-anthesis (end of booting stage toward panicle emerging)
and post-anthesis (2 weeks after flowering, during grain filling
stage), by withholding water for 6–10 days followed re-watering.
Plants were re-watered when visual stress symptoms (i.e.
le leaf rolling) appeared in the transgenic plants. To evaluate the
rate of water loss during the drought treatment, pots were
weighted daily and relative water loss was calculated.