Changes in Gibberellic Acid and Abscisic Acid Contents of Red Clover during Seed Development and Maturation.

Gibberellic acid (GA₃) and abscisic acid (ABA) are two major phytohormones that affect seed germination. While hard seed coat is the main physical dormancy form in red clover, it is not established whether physiological dormancy (hormonal balance) play any role. Therefore, the objectives of this study were to: 1) determine the amounts and the ratio of GA₃ and ABA in red clover during seed development until harvest maturity; 2) investigate the effect of these two hormones on seed germination; and 3) evaluate the effect of irrigation and foliar application of trinexapac-ethyl (TE, a plant growth hormone) at stem elongation stage on GA₃ and ABA contents. Seeds were collected from an untreated control and TE-treated plots at weekly interval after anthesis and germination tests were conducted. The GA₃ and ABA were extracted from seeds using the solid phase method and were quantified by the Liquid Chromatography and Mass Spectrometry (LC-MS) with deuterated GA₃ and ABA as internal standards. The amount of ABA was high at the early stage of seed development, and then decreased rapidly afterward, reaching the lowest level at harvest maturity. The amount of GA₃ was not significantly different during seed development until reaching harvest maturity. The ratio of ABA: GA₃ was high at the early stage of seed development, but seed germination was low. When the ratio of ABA: GA₃ decreased (approximately 2: 1) at harvest maturity, seed germination reached maximum rate at 96%. The ABA content was less in irrigated plots compared to non-irrigated plots, whereas the GA₃ content did not significantly change under irrigation treatment.  The TE application did not have significant effect on the endogenous production of GA₃ and ABA contents during seed development. These results suggest that physiological dormancy is not a substantial concern in red clover seeds.