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Abstract BackgroundModern plant breeding strategies rely on the intensive use of advanced genomic tools to expedite the development of improved crop varieties. Genomic DNA extraction from crop seeds eliminates the need to grow plants in contrast to fresh leaf tissue; however, it can still be a bottleneck due to the presence of stored compounds and the complexity of the matrix. The interaction of environmentally benign choline-based ionic liquids (ILs) with DNA offers an innovative approach to enhance the quality of extracted DNA from seeds. While prior IL-based plant DNA extraction workflows have primarily supported polymerase chain reaction (PCR) and quantitative PCR-based applications, their suitability for high-throughput sequencing (HTS) remained largely unexplored. This study explores the efficacy of IL-assisted method for genomic DNA extraction from soybean (Glycine max) seeds, addressing the limited application of ILs in HTS. ResultsThe optimized DNA extraction method, utilizing 25% (w/v) choline formate, enabled the recovery of high-purity DNA with abundant fragment sizes > 20 kb, suitable for downstream applications including PCR, whole genome amplification (WGA), simple sequence repeat (SSR) amplification, and high-throughput Illumina sequencing. The IL-method was benchmarked against a silica-binding method using cetyltrimethylammonium bromide (CTAB) and sodium dodecyl sulfate (SDS) as lysis agents using a commercial plant DNA extraction kit in terms of DNA yield, purity, abundant DNA fragment size distribution, and integrity. In addition, DNA isolated from this method demonstrated successful PCR amplification of markers from both the nuclear and plastid genomes and yielded > 99% whole genome coverage with Illumina (PE150) sequencing reads. ConclusionsThis is the first known instance of a whole genome sequence generated from DNA extracted with ILs. These findings mark a significant milestone in establishing ILs as promising alternatives to conventional methods for seed DNA extraction, with potential utility in third generation (long-read) sequencing experiments. 

Vernonia (Asteraceae: Vernonieae) is monophyletic if circumscribed to include only a North American clade of ca. 20 species. Vernonia pratensis, an endemic species of Madagascar, has been placed in the synonymy of Cyanthillium patulum or considered a distinct species in Bothriocline. In this study we characterise the pollen and cypsela of Vernonia pratensis, Bothriocline longipes, and Cyanthillium patulum with details on morphology and ultra-sculpture. The pollen grains of these species are 3-brevicolporate or 3-porate and echinolophate; in B. longipes and V. pratensis there is an inconspicuous colpus formed by interlacunar gaps (absent in C. patulum). The cypselae of B. longipes and V. pratensis have prominent ribs, broader or equally as wide as the grooves and that are fused into a shallow, apical rim; the grooves are covered by unicellular trichomes (type 1) with a striate cuticle. By contrast, the cypselae of C. patulum have ribs narrower than the grooves and not fused apically; the grooves are covered by infundibular idioblasts, and bilobed glandular trichomes at the base. Based on these morphological findings, V. pratensis is here placed in Bothriocline and named B. madagascariensis. This is the single species that occurs in Madagascar within a genus of ca. 50 species otherwise restricted to Tropical Africa and subtropical Southern Africa. Full palynological descriptions, measurements, and scanning electron microscopy (SEM) and light microscopy (LM) images are provided for the three species (B. longipes, B. madagascariensis, and C. patulum), as well as a full taxonomic description of B. madagascariensis, with a preliminary conservation status assessment, nomenclatural notes, and a discussion of possible relationships with other species of Bothriocline. 

There is a growing demand for fast and reliable plant biomolecular analyses. DNA extraction is the major bottleneck in plant nucleic acid-based applications especially due to the complexity of tissues in different plant species. Conventional methods for plant cell lysis and DNA extraction typically require extensive sample preparation processes and large quantities of sample and chemicals, elevated temperatures, and multiple sample transfer steps which pose challenges for high throughput applications. ResultsIn a prior investigation, an ionic liquid (IL)-based modified vortex-assisted matrix solid phase dispersion approach was developed using the model plant,Arabidopsis thaliana(L.) Heynh. Building upon this foundational study, the present study established a simple, rapid and efficient protocol for DNA extraction from milligram fragments of plant tissue representing a diverse range of taxa from the plant Tree of Life including 13 dicots and 4 monocots. Notably, the approach was successful in extracting DNA from a century old herbarium sample. The isolated DNA was of sufficient quality and quantity for sensitive molecular analyses such as qPCR. Two plant DNA barcoding markers, the plastidrbcLand nuclear ribosomal internal transcribed spacer (nrITS) regions were selected for DNA amplification and Sanger sequencing was conducted on PCR products of a representative dicot and monocot species. Successful qPCR amplification of the extracted DNA up to 3 weeks demonstrated that the DNA extracted using this approach remains stable at room temperature for an extended time period prior to downstream analysis. ConclusionsThe method presented here is a rapid and simple approach enabling cell lysis and DNA extraction from 1.5 mg of plant tissue across a broad range of plant taxa. Additional purification prior to DNA amplification is not required due to the compatibility of the extraction solvents with qPCR. The method has tremendous potential for applications in plant biology that require DNA, including barcoding methods for agriculture, conservation, ecology, evolution, and forensics. 

Abstract Background Plant DNA isolation and purification is a time-consuming and laborious process relative to epithelial and viral DNA sample preparation due to the cell wall. The lysis of plant cells to free intracellular DNA normally requires high temperatures, chemical surfactants, and mechanical separation of plant tissue prior to a DNA purification step. Traditional DNA purification methods also do not aid themselves towards fieldwork due to the numerous chemical and bulky equipment requirements. Results In this study, intact plant tissue was coated by hydrophobic magnetic ionic liquids (MILs) and ionic liquids (ILs) and allowed to incubate under static conditions or dispersed in a suspension buffer to facilitate cell disruption and DNA extraction. The DNA-enriched MIL or IL was successfully integrated into the qPCR buffer without inhibiting the reaction. The two aforementioned advantages of ILs and MILs allow plant DNA sample preparation to occur in one minute or less without the aid of elevated temperatures or chemical surfactants that typically inhibit enzymatic amplification methods. MIL or IL-coated plant tissue could be successfully integrated into a qPCR assay without the need for custom enzymes or manual DNA isolation/purification steps that are required for conventional methods. Conclusions The limited amount of equipment, chemicals, and time required to disrupt plant cells while simultaneously extracting DNA using MILs makes the described procedure ideal for fieldwork and lab work in low resource environments. 

Green plants play a fundamental role in ecosystems, human health, and agriculture. As de novo genomes are being generated for all known eukaryotic species as advocated by the Earth BioGenome Project, increasing genomic information on green land plants is essential. However, setting standards for the generation and storage of the complex set of genomes that characterize the green lineage of life is a major challenge for plant scientists. Such standards will need to accommodate the immense variation in green plant genome size, transposable element content, and structural complexity while enabling research into the molecular and evolutionary processes that have resulted in this enormous genomic variation. Here we provide an overview and assessment of the current state of knowledge of green plant genomes. To date fewer than 300 complete chromosome-scale genome assemblies representing fewer than 900 species have been generated across the estimated 450,000 to 500,000 species in the green plant clade. These genomes range in size from 12 Mb to 27.6 Gb and are biased toward agricultural crops with large branches of the green tree of life untouched by genomic-scale sequencing. Locating suitable tissue samples of most species of plants, especially those taxa from extreme environments, remains one of the biggest hurdles to increasing our genomic inventory. Furthermore, the annotation of plant genomes is at present undergoing intensive improvement. It is our hope that this fresh overview will help in the development of genomic quality standards for a cohesive and meaningful synthesis of green plant genomes as we scale up for the future.