A Rapid and Quantitative Method for Determining Seed Viability Using 2,3,5-Triphenyl Tetrazolium Chloride (TTC): With the Example of Wheat Seed

Abstract

Current colorimetric methods for quantitative determination of seed viability (SV) with 2,3,5-triphenyl tetrazolium chloride (TTC) have been plagued by issues of being cumbersome and time-consuming during the experimental process, slow in extraction and staining, and exhibiting inconsistent results. In this work, we introduced a new approach that combines TTC-staining with high-temperature extraction using dimethyl sulfoxide (DMSO). The optimization of the germination stage, TTC-staining method, and 1,3,5-triphenylformazan (TTF) extraction method were meticulously carried out as follows: When the majority of wheat seeds had grown the radicle, and the length of radicles was approximately equal to the seed length (24 h-germination), 2 g germinating seeds were placed into a beaker (20 mL) containing 5 mL 10 g·L−1 TTC solution. The seeds were stained with TTC in the dark at 25 °C for 1 h. Following the staining, 1 mL 1 mol·L−1 H2SO4 was added to stop the reaction for 5 min. The H2SO4 solution was then removed, and the seeds were gently rinsed with deionized water. Subsequently, the TTF produced in the seeds was extracted directly with 5 mL DMSO solution at 55 °C for 1 h. The absorbance of the extract was measured at 483 nm, and the index of SV was calculated according to a predetermined TTC calibration curve and expressed by mg TTC·g−1 (seed)·h−1. The new method has been demonstrated to be rapid, stable, and highly sensitive, as evidenced by the accurate measurement of seed viability with different aging degrees.

Effects of Aerated Drip Irrigation on the Soil Nitrogen Distribution, Crop Growth, and Yield of Chili Peppers

“…The root system was rinsed with deionized water once. After draining the water, 5 mL of acetone solution (20%) was added to the beaker to extract the 2,3,5-triphenylformazan in the root sample, and then the extent of TTC reduction was determined using a UV spectrophotometer [ 46 ].…”

“…The root system was rinsed with deionized water once. After draining the water, 5 mL of acetone solution (20%) was added to the beaker to extract the 2,3,5-triphenylformazan in the root sample, and then the extent of TTC reduction was determined using a UV spectrophotometer [46]. At the end of each reproductive period of chili peppers, soil samples were taken in layers along an S-shaped route by the soil auger method.…”

Morphological and Physiological Mechanism of Activating Insoluble Inorganic Phosphorus of Different Peanut (Arachis hypogaea L.) Varieties under Low Phosphorus

“…After 4 d of treatment with insoluble calcium phosphate, the aboveground and underground parts of the peanut seedlings were separated using scissors, and three complete roots were taken from each repeated treatment. Furthermore, 0.5 g of fresh peanut root tip samples with basically the same thickness and length were used to measure root activity via the staining method involving 2,3,5-triphenyl tetrazolium chloride (TTC) [17].…”