Extraction and determination of abscisic acid content in Arabidopsis shoots were performed as described in (Woo et al., 2011) . Briefly, 200-300 mg plant material was frozen in liquid nitrogen and ground using a Mixer-Mill (Qiagen). The homogenate was extracted with 1-2 ml ABA extraction buffer (10 mM HCl and 1% w/v polyvinyl polypyrrolidone in methanol), with continuous mixing overnight at 4 °C. After centrifugation, the supernatant was neutralized with 1 M NaOH, and ABA levels were quantified in the extract using the Phytodetek-ABA kit (AGDIA Inc.), following the manufacturer's protocol. Raw values for ABA levels were standardized on sample fresh weight and extraction volume.
Phytodetek aba kit
Manufactured by Agdia
Sourced in United States
The Phytodetek-ABA kit is a laboratory equipment designed for the detection and quantification of abscisic acid (ABA) in plant samples. The kit provides a reliable and efficient method for analyzing ABA levels, which is a crucial plant hormone involved in various physiological processes.
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5 protocols using phytodetek aba kit
1
Quantifying Abscisic Acid in Arabidopsis
2
ABA and Proline Assays in Salt-Stressed Seedlings
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For the ABA assays, eleven-day-old seedlings grown on agar plates with or without 150 mM NaCl treatment for 1 day were harvested and subsequently subjected to ABA extraction as described previously [59 (link)]. ABA was quantified using enzyme-linked immunosorbent assay (ELISA) (Phytodetek ABA kit; Agdia) in accordance with the manufacturer’s recommended protocol. The protocol for the proline assays followed the description provided in previous reports [60 (link)].
3
Quantifying Phytohormones and Sugars
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Abscisic acid content was measured as described by Lim et al [51 (link)]. Crushed seed (0.1 g) was extracted in 2ml of methanol containing 500 mg/L citric acid monohydrate and 100 mg/L butylated hydroxyl toluene, at 4°C overnight on a rotary shaker. The mixture was centrifuged at 2000 × g for 10 min, and the supernatant was transferred and dried using a speed vac (CVE-2000, EYELA). Samples were quantified using a Phytodetek-ABA kit (Agdia Inc., USA) according to the manufacturer’s instructions. Gibberellic acid (GA, MBS9310617, MyBioSource) and sugars (K-SUFRG, Megazyme, Ireland), including sucrose, D-fructose, D-glucose, were evaluated according to the manufacturer’s protocol.
4
Quantifying ABA Content in Plants
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For determination of ABA content, leaves were harvested from pepper and Arabidopsis plants treated with dehydration for 2 and 4 h and immediately frozen in liquid nitrogen. Approximately 50 mg of ground tissue were extracted overnight in 1 ml of ABA extraction buffer (methanol, containing 100 mg butylated hydroxyl toluene, 0.5 g citric acid monohydrate) at 4 °C on a rotary shaker. After centrifuged at 1500 g, the supernatant was transferred to new tube and dried using a speed vac. ABA content of each sample was quantified by using the Phytodetek-ABA kit (Agdia Inc., Elkhart, IN, USA) according to manufacturer’s instruction. ABA contents were expressed as pmol mg−1 fresh weight of the tissue.
5
Quantifying Arabidopsis ABA under Stress
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ABA content was measured as described previously (Lim and Lee, 2016 (link)). The ABA content of the rosette leaves from Arabidopsis treated with dehydration and cold stress was quantified using the Phytodetek-ABA kit (Agdia Inc., Elkhart, IN, USA), according to the manufacturer’s instructions.
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