Trans zeatin zt

Ovule culture was performed according to the method described previously by Beasley and Ting (1973) . Indole-3-acetic acid sodium salt (IAA) and gibberellic acid potassium salt (GA₃; both Sigma-Aldrich) were dissolved in distilled water and filter-sterilized to obtain stock solutions of 50 mM and 10 mM, respectively. Trans-zeatin (ZT) was dissolved in DMSO (both Sigma-Aldrich) to obtain a 50-mM stock solution. After autoclaving, plant hormones were added to the growth medium at the various experimental concentrations. The standard concentrations in the medium were 0.5 µM GA₃ and 5 µM IAA. Cotton bolls were harvested around 08.00 h at 1 d prior to anthesis and on the day of anthesis [–1 d post anthesis (DPA) and 0 DPA, respectively]. After being surface-sterilized in 75% (v/v) ethanol for 1 min and rinsed in sterile water, bolls were then soaked in 0.1% (w/v) HgCl₂ solution for 12 min for full sterilization, followed by rinsing six times with sterile water. For each treatment, 20 ovules in the middle part of each locule were excised and floated on 20 ml of liquid medium in a 100-ml flask, and then incubated in darkness at 32 °C for 3 d (for SEM) or for 15 d (for optical microscopy). Each experiment was repeated three times. An equal volume of DMSO was added to the medium as a negative control.