Evaluation and improvement of spectrophotometric assays of TTC reduction: maize (Zea mays) embryo as an example

Zhao, Pengfei; Zhu, Yuyan; Wang, Wei

doi:10.1007/s11738-009-0457-2

Cited by 19 publications

(35 citation statements)

References 30 publications

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“…Seeds were soaked in water for 4 h, cut in halves, and then stained in 0.5% TTC solution for 2 h (30°C) in the dark. The reduced TTC was extracted with acetone and assayed spectrophotometrically at 485 nm (Zhao et al 2010).…”

Section: Seed Viability Evaluationmentioning

confidence: 99%

Proteomic analysis of seed viability in maize

Liu

Wang

et al. 2010

Acta Physiol Plant

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To identify specific proteins related to maize seed viability, seeds of Zhengdan 958 (one of the highyield maize hybrids in China) were sorted based on viability evaluation with triphenyltetrazolium chloride (TTC) assay and used for comparative proteomic analysis. After TTC staining, embryos of high-viability seeds were deep red (R type), while embryos of dead seeds were white (W type). Proteomic analysis revealed that 28 protein spots identified were differently expressed significantly between R and W embryos, of which 20 were up-regulated and 8 down-regulated in R embryos. Among them were proteins involved in stress response, protein folding, and stabilization, as wells as proteins related to nutrient reservoir and metabolism. Prominently, small heat shock proteins, late embryogenesis abundant (LEA) proteins, and antioxidant enzymes were highly up-regulated, while two proteases were highly down-regulated in R embryos compared to W embryos. One of LEA proteins was EMB564, which declined in abundance during artificial aging of seeds. Our results suggested an association of EMB564 with maize seed viability. It would be of interest to use these small proteins to develop quick tests for seed quality.

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Section: Seed Viability Evaluationmentioning

confidence: 99%

Proteomic analysis of seed viability in maize

Liu

Wang

et al. 2010

Acta Physiol Plant

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“…Several seed vigor testing methods are available including the molecular test (Catusse, Strub, Job, Dorsselaer, & Job, ; Shin, Kim, & An, ), electrical conductivity test (Silva, Lopes, Marcos‐Filho, & Vieira, ), and the triphenyl tetrazolium (TTC) test (Zhao, Zhu, & Wang, ). Among them, the TTC test is a rapid and convenient method, which is used to replace traditional germination‐based assessments.…”

Section: Introductionmentioning

confidence: 99%

Optimized qualitative and quantitative methods for barley viability testing using triphenyl tetrazolium chloride staining

Ma¹,

Liu²,

Dong³

et al. 2019

Cereal Chem

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Background and objectives Barley viability is an important agronomic trait and has a direct influence on subsequent beer production. Traditional triphenyl tetrazolium chloride (TTC) test evaluating seed viability still has some drawbacks, for example, excessive instability and time consumption. In the present study, we optimized both the qualitative and quantitative TTC seed viability test methods, and established more accurate and convenient assays to evaluate barley seed viability. Findings In the qualitative evaluation method, we established the vigor index according to the proportion of staining embryos, allowing quick evaluation of both germination and mortality rates in <12 hr. In the quantitative evaluation method, we optimized TTC incubation time, extraction temperature, and solvent volume to tissue ratios making results more reliable. Finally, we developed a high‐throughput method by combining freeze drying and tissue lysing instead of the time‐consuming method of grinding on liquid nitrogen. There was a positive significant statistical correlation between germination rate and formazan content by the optimized quantitative evaluation method (R2 = 0.907; p < 0.01). Conclusions Both the optimized qualitative and quantitative TTC seed viability test methods allow a quick and reliable seed vigor evaluation for different varieties or different aged barley seeds. Significance and novelty The optimized seed vigor evaluation method can be used to effectively screen out low vigor barley seeds. The qualitative evaluation method is appropriate to quickly evaluation for a small number of samples in <12 hr. The high‐throughput quantitative evaluation method is fit for a large number of sample evaluations as much as 80 samples at the same time.

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“…However, while ethanol has the lowest vapour pressure supporting low variability during spectrophotometrics, it is also least efficient at extracting formazan (Harty et al, 1972; Zhao et al, 2010). While this inefficiency may be compensated using heat (above 80°C; Steponkus and Lanphear, 1967; Stattin and Lindström, 1999; Verleysen et al, 2004; Mikuła et al, 2006), the method also releases compounds which lead to high background and/or high ODs at 484 ηm which are not related to formazan production (Ruf and Brunner, 2003).…”

Section: Discussionmentioning

confidence: 99%

A Spectrophotometric Assay for Robust Viability Testing of Seed Batches Using 2,3,5-Triphenyl Tetrazolium Chloride: Using Hordeum vulgare L. as a Model

et al. 2017

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A comparative analysis was carried out of published methods to assess seed viability using 2,3,5-triphenyltetrazolium chloride (TTC) based assays of seed batches. The tests were carried out on seeds of barley (Hordeum vulgare cv. Optic) as a model. We established that 10% [w/v] trichloroacetic acid (TCA)/methanol is superior to the acetone and methanol-only based methods: allowing the highest recovery of formazan and the lowest background optical density (OD) readings, across seed lots comprising different ratios of viable and dead seeds. The method allowed a linear-model to accurately capture the statistically significant relationship between the quantity of formazan that could be extracted using the method we developed and the seed temperature-response, and seed viability as a function of artificially aged seed lots. Other quality control steps are defined to help ensure the assay is robust and these are reported in a Standard Operating Procedure.

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Evaluation and improvement of spectrophotometric assays of TTC reduction: maize (Zea mays) embryo as an example

Cited by 19 publications

References 30 publications

Proteomic analysis of seed viability in maize

Proteomic analysis of seed viability in maize

Optimized qualitative and quantitative methods for barley viability testing using triphenyl tetrazolium chloride staining

A Spectrophotometric Assay for Robust Viability Testing of Seed Batches Using 2,3,5-Triphenyl Tetrazolium Chloride: Using Hordeum vulgare L. as a Model

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