Fern Spores-"Ready-to-Use" Standards for Plant Genome Size Estimation Using a Flow Cytometric Approach.

Spores and pollen of plants were used as flow cytometric materials to efficiently infer genome sizes. Given this advantage, they hold great potential for various flow cytometric applications, particularly as plant genome size standards. To develop such novel standards, we investigated conditions of pretreatment (bead vortex), buffer, and reliable genome sizes of three fern spore collections-Cibotium taiwanense "Kuo4395", Sphaeropteris lepifera "Tang0001", and Alsophila metteniana "Lee s.n.". Additionally, up to 30 year-old spore collections were obtained from herbarium specimens and from samples stored at 4 °C; their spore nuclei were extracted, and the quality and quantity of these nucleus extractions through storage ages were examined. Nuclear extractions with a longer bead vortex duration or lower spore/bead ratio generally resulted in a higher recovered quantity but a lower quality or purity. For each spore standard, the protocol optimization was determined by their performance in bead vortex conditions, and a 1C genome size was further inferred by linear regression (C. taiwanense "Kuo4395" = 5.058 pg; S. lepifera "Tang0001" = 7.117 pg; and A. metteniana "Lee s.n." = 19.379 pg). Spore nucleus quality and quantity are significantly negatively correlated with storage ages. Nuclear extractions of 10-year-old refrigerated spores remained qualified as a genome size standard; however, none of the herbarium spore collections fit such criteria. Our study is the first to develop and apply dried and refrigerated spores for genome size standards. These standards are ready to use, easy to manipulate, and feature long-term storage in comparison with traditionally used standards of fresh leaves.

A dormant resource for genome size estimation in ferns: C-value inference of the Ophioglossaceae using herbarium specimen spores.

PREMISE: The great variation of genome size (C-value) across land plants is linked to various adaptative features. Flow cytometry (FCM), the standard approach to estimating C-values, relies mostly on fresh materials, performing poorly when used with herbarium materials. No fern C-value reports have been derived from herbarium specimens; however, the herbarium spores of some ferns remain highly viable for decades and are thus promising for further investigation. To explore this possibility, we evaluated herbarium spore collections of Ophioglossaceae ferns using FCM. METHODS: Flow cytometry was conducted on 24 spore samples, representing eight of the 12 genera of the Ophioglossaceae, using specimens ranging in age from 2.6 to 111 years obtained from five herbaria. RESULTS: Regardless of the genus or the source herbarium, high-quality C-value data were generated from 17 samples, with the oldest being 26 years old. Estimates of the C-values from sporophytic tissues of known ploidy did not reveal any evidence of apomixis for the species surveyed here. We also detected a pronounced genome downsizing in Sceptridium polyploids. DISCUSSION: The recent success of FCM for C-value estimation using spores provides a much more convenient method of utilizing "dry" refrigerated materials. We demonstrate here that herbarium spores of some ferns are also promising for this use, even for older specimens.

[Abundance of Toxic and Non-toxic Microcystis sp. in Lake Hongze and Its Correlation with Environmental Factors].

In the present study, the eutrophic level of 30 water samples collected from Lake hongze in August 2014 were analyzed, and the abundance of toxic and non-toxic Microcystis sp., together with their spatial distribution, was investigated by quantitative real-time PCR techniques. The results showed that the average concentrations of total nitrogen and total phosphorus were 1.63 and 0.11 mg x L(-1), respectively. The trophic state index ( TSI) ranged from 58.1 to 73.6, and the water quality was in the state of eutrophication based on TSI. Toxic Microcystis was widely distributed in Lake Hongze, and its abundance varied sharply, from 1. 13 x 10(4) to 3.51 x 10(6) copies x mL(-1), and the abundance of total Microcystis ranged from 1.06 x 10(5) to 1.10 x 10(7) copies x m(-1), meanwhile, the proportion of toxic Microcystis in the total Microcystis ranged from 8.5% to 38.5%, with the average value of 23.6%. Correlation analysis indicated that there was a significant positive correlation among total Mirocystis, toxic Microcystis and the toxic proportion (P < 0.01). The abundance of total and toxic Microcystis was significantly positively correlated to chlorophyll a ( Chl-a) concentrations and TSI (P < 0.01), but was negatively correlated to transparency (SD) (P < 0.01). The ratio of toxic Microcystis to total Microcystis was significantly positively correlated to Chl-a, TN, TP and TSI (P < 0.01), but significantly negatively correlated to the ratio of TN to TP and SD (P < 0.01). Therefore, reducing total nitrogen and phosphorus concentrations could not only lower the eutrophication level of Lake Hongze, but also inhibit the competition advantage of the toxic Microcystis over non-toxic Microcystis.