Apomixis, clonal reproduction through seed, is divided into sporophytic (nucellar embryony) and gametophytic apomixis. This book focuses on the latter type. Gametophytic apomixis, while rare, is taxonomically widespread, notably among grasses and Asteraceae. Detection methods have advanced significantly, employing flow cytometry and advanced microscopy techniques. The genetic, molecular, and developmental mechanisms behind apomixis involve modifications preventing meiosis and enabling parthenogenesis. Understanding these mechanisms and applying approaches such as forward genetics and mutagenesis holds an important potential for agricultural innovation. Numerous genetic, ecological, and evolutionary questions remain open, highlighting apomixis as an important field for ongoing scientific research.

Apomixis-clonal reproduction through seeds-is an alternative reproductive strategy that takes place in less than 0.1% of plant species and has evolved independently in diverse plant lineages. To date, the genetic basis of apomixis has been unraveled genetically in only a few genera. The identification of causal apomixis genes is technically challenging, as apomictic species are typically polyploid and the genetic loci associated with apomixis are often in low-recombination regions limiting conventional fine-mapping. In triploid apomictic dandelion (Taraxacum officinale), after conventional genetic mapping, deletion mapping, and complete apomixis loci haplotype assembly, we made use of targeted mutagenesis using CRISPR/Cas9 technology to identify the Taraxacum officinale PARTHENOGENESIS (ToPAR) gene that is responsible for embryogenesis in the absence of fertilization. Here, we report the methods used to clone the ToPAR gene by targeted mutagenesis and we expect that the general principles could be applied in other systems to identify novel apomixis genes.

Embryogenesis in plants is a process that not only occurs after fertilization but can also be initiated in its absence. This form of embryogenesis is known as parthenogenesis, and it is an essential part of a process called apomixis. Apomixis has long been seen as the holy grail of plant breeding because it produces clonal seeds. Therefore, engineering (single components of) apomixis remains a highly desired research goal. Over the last few years, significant progress has been made toward understanding and engineering both complete synthetic apomixis system and individual components of apomixis, such as parthenogenesis, in crops. Characterization of candidate parthenogenesis genes involves thorough phenotypic analysis and microscopic observation of developing embryos, which are, however, not readily accessible because they are embedded in several integumental layers comprising the protective seed coat. Here, we provide a detailed approach for detecting parthenogenesis and we further demonstrate its application to a sexual crop species, i.e., lettuce. We describe an emasculation technique by decapitation, embryo sac isolation, imaging of cleared embryos, and ploidy analysis of embryo sacs utilizing flow cytometry.

This chapter describes the erratic course of the research journey of our research group into the elucidation of the genetic basis of apomixis in the common dandelion, Taraxacum officinale s. l. Research is rarely straightforward; sometimes, it comes to a standstill, requiring a shift in approach. Our journey took years to complete, starting with the theoretical genetic models and culminating in the cloning of the parthenogenesis gene. Along the way, we employed a suite of genetic, cell biological, and molecular techniques.

The female germline precursor cell, known as the megaspore mother cell (MMC), differentiates and develops within the ovule through a complex genetic network, including post-transcriptional and translational regulation mediated by miRNA. A key player in restricting MMC development to a single cell within the nucellus is the AUXIN RESPONSE FACTOR 3 (ARF3). Here, we describe the role of the VACUOLAR PROTEIN SORTING-ASSOCIATED 13 (VPS13) in post-transcriptional regulation of ARF3. VPS13 is a conserved protein found across all eukaryotes, implicated in membrane dynamics, organelle junctions, and autophagy. We show that in Arabidopsis, the knockout mutation of VPS13 results in the deregulation of ARF3, most likely due to a reduction of tasiR-ARF production. Consequently, vps13 mutant ovules exhibit phenotypes similar to those caused by ARF3 misregulation, such as the formation of multiple MMC-like cells and defects in megasporogenesis. VPS13 co-localizes and interacts with the SUPPRESSOR OF GENE SILENCING 3 (SGS3). Based on our findings, we propose an uncharacterized potential role for VPS13 in the homeostasis of membrane-associated siRNA bodies, essential for tasiR-ARF biogenesis. Thus, it adds an additional component to the intricate network governing female germline initiation and progression, while also providing new insights into siRNA body homeostasis and tasiR-ARF pathways. Furthermore, the evolutionary conservation of VPS13 suggests its broader relevance, potentially extending to the understanding of human diseases linked to mutations in its orthologues.

Apomictic plants are able to produce clonal seeds. This reproductive system allows the one-step fixation of any valuable trait for subsequent generations and would pave the way for a revolution in the agricultural system. Despite this, the introduction of apomixis in sexually reproducing crops has been hampered due to the difficulty in characterizing its genetic regulation. In this study, we described the high-resolution characterization of apomeiosis in the apomictic model species Erigeron annuus, Chondrilla juncea, and Taraxacum officinale. We showed that apomeiosis differs from meiosis in a few critical steps, including homologous chromosome synapsis and segregation during meiosis I. We then compared megasporogenesis in three T. officinale genetic backgrounds, showing that diplospory is superimposed on the sexual pathway without severely altering the expression of crucial meiotic genes. Our findings will contribute to the identification of pivotal players controlling this intriguing asexual reproductive strategy.

According to the revisionist interpretation of Mendel's pea crosses, his primary aim was not to study the inheritance of traits. Instead, he was interested in the question raised by Linnaeus as to whether new species could arise from the hybridization of existing species. The genetic interpretation is therefore seen as ahistorical by the revisionists. This view goes back to the 1979 article "Mendel no Mendelian?" by the historian of science R.C. Olby. A closer analysis shows that Olby implicitly assumed Mendel adhered to the unusual strictest species definition for Pisum. However, we argue that Mendel only mentions the hypothetical application of this strict definition in his 1866 paper. Like most of his contemporaries, Mendel accepted variation within species where the differences between varieties and species were a matter of degree. After researching variable hybrids in peas (Pisum; 1854-1863), Mendel also studied constant hybrids in hawkweeds (Hieracium; 1866-1873), which he considered to be new species. There is no debate about the latter, but the matter becomes muddled because Olby lumps Pisum and Hieracium together, despite their having completely different reproduction systems. Based on newly discovered historical sources, we also dispute several other assumptions made by Olby. We do not consider Olby's claim that Mendel conducted the Pisum experiments to investigate species multiplication to be tenable.

Apomixis is considered a potentially revolutionary tool to generate high-quality food at a lower cost and shorter developmental time due to clonal seed production through apomeiosis and parthenogenesis. In the diplosporous type of apomixis, meiotic recombination and reduction are circumvented either by avoiding or failing meiosis or by a mitotic-like division. Here, we review the literature on diplospory, from early cytological studies dating back to the late 19th century to recent genetic findings. We discuss diplosporous developmental mechanisms, including their inheritance. Furthermore, we compare the strategies adopted to isolate the genes controlling diplospory with those to produce mutants forming unreduced gametes. Nowadays, the dramatically improved technologies of long-read sequencing and targeted CRISPR/Cas mutagenesis justify the expectation that natural diplospory genes will soon be identified. Their identification will answer questions such as how the apomictic phenotype can be superimposed upon the sexual pathway and how diplospory genes have evolved. This knowledge will contribute to the application of apomixis in agriculture.

We describe both the terminology and use of symbols introduced by Mendel in his 1866 paper and discuss some misconceptions concerning their interpretation.

The pharmacokinetics and metabolism of reboxetine, a selective noradrenaline reuptake inhibitor, in humans and animal models are reviewed here. Reboxetine has potent antidepressant activity, low affinity for alpha-adrenergic and muscarinic receptors and low toxicity in animals. It is a mixture of (R,R) and (S,S) enantiomer, the latter being more potent but no qualitative differences in pharmacodynamic properties are observed between the two. Humans rapidly absorb reboxetine (tmax about 2 h) with a terminal half-life of elimination (t½) of 13 h, allowing twice-daily administration. Animal models also rapidly absorb reboxetine (tmax 0.5–2 h) but t½ was 1–2 h. Food does not affect bioavailability. There were no major inter-species differences in the metabolic profile of reboxetine. Elimination is principally renal in humans and monkeys. Reboxetine has linear pharmacokinetics in young, healthy males for single doses of 1–5 mg and in elderly, female depressed patients (up to 4 mg b.i.d.). Multiple dosing, gender or liver insufficiency had no significant effects on the pharmacokinetics. Elderly (particularly frail elderly) patients and patients with severe renal impairment may need dose reduction. Reboxetine shows no clinically relevant interaction with lorazepam and has no inhibitory effects on the major enzymes involved in drug metabolism. It may be possible to use reboxetine in combination with monoamine oxidase inhibitors as it has no inhibitory effect on this enzyme; in addition, it may protect patients against tyramine-induced reactions. In conclusion, reboxetine seems to be an antidepressant with negligible interference with the pharmacokinetics of other drugs thus fewer drug–drug interactions are expected.

Apomixis, the clonal formation of seeds, is a rare yet widely distributed trait in flowering plants. We have isolated the PARTHENOGENESIS (PAR) gene from apomictic dandelion that triggers embryo development in unfertilized egg cells. PAR encodes a K2-2 zinc finger, EAR-domain protein. Unlike the recessive sexual alleles, the dominant PAR allele is expressed in egg cells and has a miniature inverted-repeat transposable element (MITE) transposon insertion in the promoter. The MITE-containing promoter can invoke a homologous gene from sexual lettuce to complement dandelion LOSS OF PARTHENOGENESIS mutants. A similar MITE is also present in the promoter of the PAR gene in apomictic forms of hawkweed, suggesting a case of parallel evolution. Heterologous expression of dandelion PAR in lettuce egg cells induced haploid embryo-like structures in the absence of fertilization. Sexual PAR alleles are expressed in pollen, suggesting that the gene product releases a block on embryogenesis after fertilization in sexual species while in apomictic species PAR expression triggers embryogenesis in the absence of fertilization.

There are few historical records concerning Gregor Johann Mendel and his work, so theories abound concerning his motivation. These theories range from Fisher’s view that Mendel was testing a fully formed previous theory of inheritance to Olby’s view that Mendel was not interested in inheritance at all, whereas textbooks often state his motivation was to understand inheritance. In this Perspective, we review current ideas about how Mendel arrived at his discoveries and then discuss an alternative scenario based on recently discovered historical sources that support the suggestion that Mendel’s fundamental research on the inheritance of traits emerged from an applied plant breeding program. Mendel recognized the importance of the new cell theory; understanding of the formation of reproductive cells and the process of fertilization explained his segregation ratios. This interest was probably encouraged by his friendship with Johann Nave, whose untimely death preceded Mendel’s first 1865 lecture by a few months. This year is the 200th anniversary of Mendel’s birth, presenting a timely opportunity to revisit the events in his life that led him to undertake his seminal research. We review existing ideas on how Mendel made his discoveries, before presenting more recent evidence.

Apomixis in the common dandelion (Taraxacum officinale) consists of three developmental components: diplospory (apomeiosis), parthenogenesis, and autonomous endosperm development. The genetic basis of diplospory, which is inherited as a single dominant factor, has been previously elucidated. To uncover the genetic basis of the remaining components, a cross between a diploid sexual seed parent and a triploid apomictic pollen donor was made. The resulting 95 triploid progeny plants were genotyped with co-dominant simple-sequence repeat (SSR) markers and phenotyped for apomixis as a whole and for the individual apomixis components using Nomarski Differential Interference Contrast (DIC) microscopy of cleared ovules and seed flow cytometry. From this, a new SSR marker allele was discovered that was closely linked to parthenogenesis and unlinked to diplospory. The segregation of apomixis as a whole does not differ significantly from a three-locus model, with diplospory and parthenogenesis segregating as unlinked dominant loci. Autonomous endosperm is regularly present without parthenogenesis, suggesting that the parthenogenesis locus does not also control endosperm formation. However, the high recovery of autonomous endosperm is inconsistent with this phenotype segregating as the third dominant locus. These results highlight the genetic complexity underlying apomixis in the dandelion and underline the challenge of introducing autonomous apomixis into sexual crops.

Taraxacum koksaghyz Rodin, a dandelion from the steppes of south-eastern Kazakhstan, has been known for long time as potential rubber producer, as a temperate region alternative to the tropical rubber tree Hevea brasiliensis Müll. Arg.. In this work, we evaluate Taraxacum bicorne Dahlst. (wild populations), a closely related congener of T. koksaghyz. The taxonomy of T. bicorne is reviewed, population genetic characteristics are analysed by Simple Sequence Repeat markers, and rubber biosynthesis genes are identified by Kompetitive Allele Specific PCR. Rubber content of T. bicorne plants collected in wild is measured using a modified extraction method to minimize amount of material. Taraxacum bicorne is shown to be an outcrossing sexual diploid. Its rubber content is about half of that of T. koksaghyz (~ 3.2% vs. ~ 7.2% of root dry weight), but T. bicorne could be used as a potential rubber source or as a source of germplasm for production of hybrids to improve rubber yield in various environments, because T. bicorne has considerable genetic variation, wider geographical range than T. koksaghyz and grows in different environments.

What is rubber and where does it come from? Rubber is a natural product produced by plants and is present in many of the goods used in our daily lives.Rubber has had an important role in human history, throughout the development of human civilizations. It still plays an important role, and that is why we need to search for new rubber sources. Nowadays, 99% of the natural rubber we use is extracted from a tree called Hevea brasiliensis. In this article, we give some details about the best alternative rubber sources currently available.

A controversy arose over Mendel’s pea crossing experiments after the statistician R.A. Fisher proposed how these may have been performed and criticised Mendel’s interpretation of his data. Here we re-examine Mendel’s experiments and investigate Fisher’s statistical criticisms of bias. We describe pea varieties available in Mendel’s time and show that these could readily provide all the material Mendel needed for his experiments; the characters he chose to follow were clearly described in catalogues at the time. The combination of character states available in these varieties, together with Eichling’s report of crosses Mendel performed, suggest that two of his F3 progeny test experiments may have involved the same F2 population, and therefore that these data should not be treated as independent variables in statistical analysis of Mendel’s data. A comprehensive re-examination of Mendel’s segregation ratios does not support previous suggestions that they differ remarkably from expectation. The χ 2 values for his segregation ratios sum to a value close to the expectation and there is no deficiency of extreme segregation ratios. Overall the χ values for Mendel’s segregation ratios deviate slightly from the standard normal distribution; this is probably because of the variance associated with phenotypic rather than genotypic ratios and because Mendel excluded some data sets with small numbers of progeny, where he noted the ratios “deviate not insignificantly” from expectation.

Despite the fact that Gregor Mendel is generally respected as the founder of genetics, little is known about the origin of and motivation for his revolutionary work. No primary sources are known that discuss his work during the period of his pea crossing experiments. Here, we report on two previously unknown interconnected local newspaper articles about Mendel's work that predate his famous Pisum lectures by 4 years. These articles describe Mendel as a plant breeder and a horticulturist. We argue that Mendel's initial interests concerned crop improvement, but that with time he became more interested in fundamental questions about inheritance, fertilization, and natural hybridization.

'DRIVE4EU - Dandelion Rubber and Inulin Valorization and Exploitation for Europe', a demonstration project, aims at the development of a value chain for natural rubber and inulin from Rubber dandelions. The objective of the project is to set up a new European chain for the production and processing of natural rubber. This will enable the EU to become less dependent on the import of natural rubber and at the same time to respond to the threat of a global rubber shortage. The viability of using Rubber dandelions for rubber and inulin for bioplastics (PEF - Polyethylene Furanoate) production depends on the sustainability of this new value chain. Within the project an environmental assessment using the methodology of Life Cycle Assessment (LCA) is performed. The aim is to identify, quantify and assess the most important environmental impacts and benefits of rubber and inulin from Rubber dandelion based on the whole value chain. Within the LCA scientific environmental indicators (e.g. global warming potential, cumulated primary energy demand, land use, water use, and acidification) will be used to guide the development of the DRIVE4EU value chain to realize the highest possible sustainability in comparison to a substituted reference system (natural rubber from Hevea tree and PET from fossil resources). The combination of natural rubber and inulin makes Rubber dandelion very interesting as a production platform.

Sexual reproduction is the predominant mode of reproduction in flowering plants, however a small but significant number of flowering plant species can produce asexual seeds, a type of reproduction known as apomixis. Apomixis can be divided into sporophytic and gametophytic apomixis. In the case of sporophytic apomixis one or more somatic embryos are formed, in addition to the sexual embryo. Gametophytic apomixis involves modification of sexual development by the avoidance of meiotic reduction, and the subsequent parthenogenetic development of the egg cell into an embryo. Gametophytic apomixis is closely associated with polyploidy and controlled by dominant genes which suppress the sexual pathway. Epigenetic pathways may also be involved in the control of the sexual and apomictic development. Apomixis does not occur in major crops, but introduction of apomixis in sexual crop species has great potential for plant breeding and seed production.

'DRIVE4EU - Dandelion Rubber and Inulin Valorization and Exploitation for Europe', a demonstration project, aims at the development of a value chain for natural rubber and inulin from Rubber dandelions. The objective of the project is to set up a new European chain for the production and processing of natural rubber. This will enable the EU to become less dependent on the import of natural rubber and at the same time to respond to the threat of a global rubber shortage. The viability of using Rubber dandelions for rubber and inulin production depends on the sustainability of this new value chain. The results of a general economic assessment shows that the total costs over the whole value chain are dominated by the costs for cultivation and harvesting and the cost for biorefining. The combination of natural rubber and inulin makes Rubber dandelion very interesting as a production platform.