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Light promoted brown staining of protoplasm by Ag+ is ideal to test wheat pollen viability rapidly.

ABSTRACT: Pollen viability is crucial for wheat breeding programs. The unique potential of the protoplasm of live cells to turn brown due to the synthesis of silver nanoparticles (AgNPs) through rapid photoreduction of Ag+, was exploited for testing wheat pollen viability. Ag+-viability test medium (consisting of 0.5 mM AgNO3 and 300 mM KNO3) incubated with wheat pollen turned brown within 2 min under intense light (~600 ?mol photon flux density m-2s-1), but not in dark. The brown medium displayed AgNPs-specific surface plasmon resonance band in its absorption spectrum. Light microscopic studies showed the presence of uniformly stained brown protoplasm in viable pollen incubated with Ag+-viability medium in the presence of light. Investigations with transmission electron microscope coupled with energy dispersive X-ray established the presence of distinct 5-35 nm NPs composed of Ag. Powder X-ray diffraction analysis revealed that AgNPs were crystalline and biphasic composed of Ag0 and Ag2O. Conversely, non-viable pollen and heat-killed pollen did not turn brown on incubation with Ag+-medium in light. We believe that the viable wheat pollen turn brown rapidly by bio-transforming Ag+ to AgNPs through photoreduction. Our findings furnish a novel simplest and rapid method for testing wheat pollen viability.

SUBMITTER: Biswas A

PROVIDER: S-EPMC7743923 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Light promoted brown staining of protoplasm by Ag+ is ideal to test wheat pollen viability rapidly.

Biswas Abhishek A Divya Subramaniyan S Sharmila Peddisetty P Pardha-Saradhi Peddisetty P

PloS one 20201216 12

Pollen viability is crucial for wheat breeding programs. The unique potential of the protoplasm of live cells to turn brown due to the synthesis of silver nanoparticles (AgNPs) through rapid photoreduction of Ag+, was exploited for testing wheat pollen viability. Ag+-viability test medium (consisting of 0.5 mM AgNO3 and 300 mM KNO3) incubated with wheat pollen turned brown within 2 min under intense light (~600 μmol photon flux density m-2s-1), but not in dark. The brown medium displayed AgNPs-s ...[more]

PMID: 33326483

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Project description:Wheat sheds tricellular short-lived pollen at maturity. The identification of viable pollen required for high seed set is important for breeders and conservators. The present study aims to evaluate and improve pollen viability tests and to identify factors influencing viability of pollen. In fresh wheat pollen, sucrose was the most abundant soluble sugar (90%). Raffinose was present in minor amounts. However, the analyses of pollen tube growth on 112 liquid and 45 solid media revealed that solid medium with 594 mM raffinose, 0.81 mM H3BO3, 2.04 mM CaCl2 at pH5.8 showed highest pollen germination. Partly or complete substitution of raffinose by sucrose, maltose, or sorbitol reduced in vitro germination of the pollen assuming a higher metabolic efficiency or antioxidant activity of raffinose. In vitro pollen germination varied between 26 lines (P < 0.001); between winter (15.3 ± 8.5%) and spring types (30.2 ± 13.3%) and was highest for the spring wheat TRI 2443 (50.1 ± 20.0%). Alexander staining failed to discriminate between viable, fresh pollen, and non-viable pollen inactivated by ambient storage for >60 min. Viability of fresh wheat pollen assessed by fluorescein diacetate (FDA) staining and impedance flow (IF) cytometry was 79.2 ± 4.2% and 88.1 ± 2.7%, respectively; and, when non-viable, stored pollen was additionally tested, it correlated at r = 0.54 (P < 0.05) and r = 0.67 (P < 0.001) with in vitro germination, respectively. When fresh pollen was used to assess the pollen viability of 19 wheat, 25 rye, 11 barley, and 4 maize lines, correlations were absent and in vitro germination was lower for rye (11.7 ± 8.5%), barley (6.8 ± 4.3%), and maize (2.1 ± 1.8%) pollen compared to wheat. Concluding, FDA staining and IF cytometry are used for a range of pollen species, whereas media for in vitro pollen germination require specific adaptations; in wheat, a solid medium with raffinose was chosen. On adapted media, the pollen tube growth can be exactly analyzed whereas results achieved by FDA staining and IF cytometry are higher and may overestimate pollen tube growth. Hence, as the exact viability and fertilization potential of a larger pollen batch remains elusive, a combination of pollen viability tests may provide reasonable indications of the ability of pollen to germinate and grow.

Project description:Terminal heat stress during reproductive stage in wheat (Triticum aestivum L.) causes pollen grain sterility and has a drastic impact on wheat crop production. Finding genotypes with high pollen viability under heat stress is crucial to cope with the impact of climate change through developing heat-tolerant cultivars. To assess the effect of terminal heat stress on pollen viability in a panel of spring wheat genotypes (N = 200), RCBD (randomized complete block design) field trials were conducted under normal and heat stress conditions for two consecutive years (2020-2021 and 2021-2022). Analysis of variance showed significant variation in genotypes, treatments, and genotype × treatment interaction. Fifty and 46 genotypes were categorized as heat tolerant (HSI pv < 0.5) in the first and second year, respectively. Twelve genotypes, namely, Chenab-70, Pari-73, Pak-81, MH-21, Punjab-76, NIFA-Aman, NUWYT-63, Swabi-1, Nisnan-21, Frontana, Amin-2000, and Pirsabak-2004, were found to be heat tolerant across the years. The violin plot displayed a trend of improvement in heat tolerance (HSI pv < 0.5) over the period of time in many modern wheat varieties. However, some modern wheat varieties released after 2001 such as Janbaz-09 (57%), Ghazi-2019 (57%), and Sindhu-16 (43%) had very low pollen viability under heat stress conditions. The results of phenotypic coefficient of variance (PCV%), genotypic coefficient of variance (GCV%), broad sense heritability (h2 bs), and genetic advance (GA) suggested the major contribution of genetic factors in controlling pollen viability trait. Higher values of h2 bs and GA under heat stress conditions suggested pollen viability as a heat tolerance trait controlled by additive genetic effects. Taken together, these results suggested pollen viability as a useful trait for selection in early generations under elevated temperatures. The genotypes identified as heat tolerant in both years can be used as genetic resources for breeding cultivars with higher pollen viability under elevated temperature conditions.

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Light promoted brown staining of protoplasm by Ag+ is ideal to test wheat pollen viability rapidly.

Publications

Light promoted brown staining of protoplasm by Ag+ is ideal to test wheat pollen viability rapidly.

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