Androgenetic Research Articles

Ozone stress-induced DNA methylation variations and their transgenerational inheritance in foxtail millet.

Elevated near-surface ozone (O3) concentrations have surpassed the tolerance limits of plants, significantly impacting crop growth and yield. To mitigate ozone pollution, plants must evolve a rapid and effective defense mechanism to alleviate ozone-induced damage. DNA methylation, as one of the most crucial epigenetic modifications, plays a pivotal role in maintaining gene stability, regulating gene expression, and enhancing plant resilience to environmental stressors. However, the epigenetic response of plants to O3 stress, particularly DNA methylation variations and their intergenerational transmission, remains poorly understood. This study aims to explore the epigenetic mechanisms underlying plant responses to ozone stress across generations and to identify potential epigenetic modification sites or genes crucial in response to ozone stress. Using Open Top Chambers (OTCs), we simulated ozone conditions and subjected foxtail millet to continuous ozone stress at 200 nmol mol-1 for two consecutive generations (S0 and S1). Results revealed that under high-concentration ozone stress, foxtail millet leaves exhibited symptoms ranging from yellowing and curling to desiccation, but the damage in the S1 generation was not more severe than that in the S0 generation. Methylation Sensitive Amplified Polymorphism (MSAP) analysis of the two generations indicated that ozone stress-induced methylation variations ranging from 10.82% to 13.59%, with demethylation events ranged from 0.52% to 5.58%, while hypermethylation occurred between 0.35% and 2.76%. Reproductive growth stages were more sensitive to ozone than vegetative stages. Notably, the S1 generation exhibited widespread demethylation variations, primarily at CNG sites, compared to S0 under similar stress conditions. The inheritance pattern between S0 and S1 generations was mainly of the A-A-B-A type. By recovering and sequencing methylation variant bands, we identified six stress-related differential amplification sequences, implicating these variants in various biological processes. These findings underscore the potential significance of DNA methylation variations as a critical mechanism in plants' response to ozone stress, providing theoretical insights and references for a comprehensive understanding of plant adaptation mechanisms to ozone stress and the epigenetic role of DNA methylation in abiotic stress regulation.

Seasonal hypoxia enhances sediment iron-bound phosphorus release in a subtropical river reservoir

Dams worldwide commonly accelerate the eutrophication of reservoirs. While the seasonal hypoxia in deep reservoirs is widely acknowledged, there is limited research on its impact on benthic phosphorus (P) cycling and P fraction release from the reservoir sediments. Here we show that seasonal hypoxia enhances sediment P release and P fluxes at the sediment-water interface (SWI) which might alter P dynamics in deep reservoirs. We conducted a detailed measurement of sediment P fractions through the SEDEX approach, combined with a labile P gradient analysis using the diffusive gradients in thin films (DGT) technique to understand P cycling patterns in sediments during the transition period from spring (oxic) to late summer (hypoxic) conditions. The sediment P pool was predominantly composed of iron-bound phosphorus (Fe−P, 76–80 %), primarily due to the widespread occurrence of lateritic red soil (rich in Fe2O3/MnO2) in subtropical areas. More organic-P was observed in summer compared to spring. A significant increase in labile P occurred at the depth of 0–4 cm and 0–1 cm in spring and summer, respectively, where sediment P release was primarily governed by the reduction of Fe−P and the generation of S2−. A higher apparent fluxes of phosphate across the SWI were observed in summer characterized by higher temperature and lower oxygen levels. The current results suggest that seasonal hypoxia was a crucial factor affecting P cycling and diffusion in deep reservoirs. These findings present important implications for the ecology and management of the watershed-coast ecosystem.

The selection index of S3 corn convergent breeding population based on multivariate analysis

Abstract. Makmur, Farid M, Ala A, Mandja K, Anshori MF, Fadhilah AN. 2024. The selection index of S3 corn convergent breeding population based on multivariate analysis. Biodiversitas 25: 1097-1103. Corn is one of the most important and strategic food crops in the world, including Indonesia. Increasing corn production through a plant breeding program by assembling hybrid varieties begins with developing a convergent breeding population and index selection. The research aims to identify genetic variance and the effectiveness of selection indices in selecting high-yield S3 cross populations. The research was carried out in Rea Village, Binuang Sub-District, Polewali Mandar District, West Sulawesi from July to November 2022. The research used an augmented design with four blocks. Sixty-six lines were derived from 4 double cross populations and 8 populations of the S3 generation multiple-cross population, as well as 9 check varieties (NK7328, Pioneer 36, Bisi 18, Bisi 2, NK99, Nasa 29, JH45, Bisi 9, and NK212) were used in this study. The results indicate that the length of cobs (5.98;73.59%), length of seed cobs (7.06; 79.46%), weight of 100 seeds (7.75; 89.30%), and weight of seeds per cob (7.84; 65.59%) have moderate genetic variance and high heritability. The characteristics of stem diameter, yield percentage, and weight of seed per cob were effectively used in the selection index formulation. Based on the index selection, there were 14 potential corn lines with a good performance, recommended for the next hybridization stage.

Is apomixis occurring in walnut (Juglans regia L.)? New data from progeny molecular tests and cytological investigations shed light on its reproductive system.

Persian walnut (Juglans regia) is an economically important nut fruit species cultivated worldwide for its nutritious kernel and timber quality wood. Walnut trees are mostly hetero-dichogamous and, depending on the genotype, some cultivars are protogynous, while others are protandrous. Although selfing is possible when male and female blooms overlap, the dichogamy of the species promotes outcrossing. In addition to sexual reproduction, some reports indicate that elements of apomixis may occur in commercial orchards of walnut varieties and in the last two decades, nut production by apomixis has been reported in walnut. However, there are no reliable studies on the occurrence of apomictic reproduction based on cytoembryological observations and/or molecular marker-progeny tests. This study addresses the combined use of molecular and cytological analyses to gain new insights into the population genetics and reproduction systems of J. regia. We systematically analyzed the reproductive origin of individual progeny plants from 8 different cultivated walnut genotypes using microsatellite genotyping and carried out cytohistological investigations of 5 cultivated walnut genotypes arising seed sets from isolated flowers, to shed light on the mode of reproduction. These cytometric and genotyping analyses did not support any asexual mode of reproduction or asexual propagation by seed and all individuals studied were identified as zygotic plants produced by crossing. Likewise, the cytological findings did not confirm completely the first component of apomixis, namely apomeiosis. On the other hand, according to histological evidence, adventitious embryony seems to take place at low frequency. Overall, our findings suggest that the occurrence of gametophytic apomixis is unlikely in J. regia, but sporophytic apomixis cannot be completely ruled out.

A Set of Yellow Mustard (Sinapis alba L.) Germplasm with Polycotyledony

A world collection of 132 yellow mustard (Sinapis alba L.) accessions was characterized in a greenhouse to identify germplasm with polycotyledony and to assess the genetic segregation of tricot and tetracot seedlings in selfed S1 and S2 generations. The effort identified a set of 46 yellow mustard accessions with frequent occurrences of polycotyledony. The revealed genetic segregations seemed to suggest the development of tricot and tetracot seedlings in yellow mustard was largely controlled by a combination of genes at multiple diallelic loci. The identified tricot germplasm can be used to facilitate the genetic and/or genomic analysis of polycotyledonous germplasm for a better understanding of genetic and developmental mechanisms conditioning polycotyledony and the development of yellow mustard lines for explorable tricot breeding.

Promoter insertion leads to polyembryony in mango — a case of convergent evolution with citrus

Abstract Sexual reproduction in plants is the main pathway for creating new genetic combinations in modern agriculture. In heterozygous plants, after the identification of a plant with desired traits, vegetative propagation (cloning) is the primary path to create genetically uniform plants. Another natural plant mechanism that creates genetically uniform plants (clones) is apomixis. In fruit crops like citrus and mango, sporophytic apomixis results in polyembryony, where seeds contain multiple embryos, one of which is sexually originated and the others are vegetative clones of the parent mother tree. Utilizing the mango genome and genetic analysis of a diverse germplasm collection, we identified MiRWP as the gene that causes polyembryony in mango. There is a strong correlation between a specific insertion in the gene’s promoter region and altered expression in flowers and developing fruitlets, inducing multiple embryos. The MiRWP gene is an ortholog of CitRWP that causes polyembryony in citrus. Based on the data, we speculate that promoter insertion events, which occurred independently in citrus and mango, induced nucellar embryogenesis. The results suggest convergent evolution of polyembryony in the two species. Further work is required to demonstrate the utility of these genes (mango and citrus) in other biological systems as a tool for the clonal production of other crops.

Electrosynthesis and characterization of alloyed CdSxSe1−x ternary quantum dots

CdS, CdSe and CdSxSe1−x (x = 0.15, 0.35, 0.5, 0.65, 0.85) ternary quantum dots (QDs) were electrosynthesized in a cavity cell and aqueous medium, using 3-mercaptopropionic acid (MPA) as stabilizer. The electrochemical procedure was carried out by a paired electrolysis, where elemental sulfur and selenium were reduced in a graphite powder macroelectrode while a sacrificial cadmium rod was oxidized, giving the simultaneous generation of S2-, Se2- and Cd2+ ions for efficient production of the ternary QDs, in the S2-/Se2- ratio of choice. The CdSxSe1−x QDs were characterized by XRD, HRTEM, UV-Vis absorption and emission spectroscopies, and voltammetric analyses. XRD analysis showed lattice parameters of the cubic CdSxSe1−x nanoalloys with a linear drop tendency when the sulfide molar ratio increased in the crystalline structure. HRTEM analysis was used to determine nanoparticle sizes: 3.94 ± 0.70 nm (CdS), 4.23 ± 1.00 nm (CdS0.5Se0.5), and 4.12 ± 0.78 nm (CdSe), with interplanar distances of 0.348 nm, 0.341 nm, and 0.339 nm, respectively. The band gap energy (Eg opt) of the CdSxSe1−x QDs were determined by UV-Vis absorption spectroscopy: 2.42, 2.47, 2.53, 2.66, 2.63, 2.73 and 2.84 eV, respectively, showing a linear increasing according to the S2-/Se2- ratio. The same behavior was observed for the Eg Elect determined by cyclic and linear voltammetry analysis, presenting a nanoalloy nature of the ternary QDs electrosynthesized.

New insights on the evolution of nucleolar dominance in newly resynthesized hexaploid wheat Triticum zhukovskyi.

Nucleolar dominance (ND) is a widespread epigenetic phenomenon in hybridizations where nucleolus transcription fails at the nucleolus organizer region (NOR). However, the dynamics of NORs during the formation of Triticum zhukovskyi (GGAu Au Am Am ), another evolutionary branch of allohexaploid wheat, remains poorly understood. Here, we elucidated genetic and epigenetic changes occurring at the NOR loci within the Am , G, and D subgenomes during allopolyploidization by synthesizing hexaploid wheat GGAu Au Am Am and GGAu Au DD. In T. zhukovskyi, Au genome NORs from T. timopheevii (GGAu Au ) were lost, while the second incoming NORs from T. monococcum (Am Am ) were retained. Analysis of the synthesized T. zhukovskyi revealed that rRNA genes from the Am genome were silenced in F1 hybrids (GAu Am ) and remained inactive after genome doubling and subsequent self-pollinations. We observed increased DNA methylation accompanying the inactivation of NORs in the Am genome and found that silencing of NORs in the S1 generation could be reversed by a cytidine methylase inhibitor. Our findings provide insights into the ND process during the evolutionary period of T. zhukovskyi and highlight that inactive rDNA units may serve as a 'first reserve' in the form of R-loops, contributing to the successful evolution of T. zhukovskyi.

Genome-wide identification of the C2H2-Zinc finger gene family and functional validation of CsZFP7 in citrus nucellar embryogenesis.

Genome-wide identification of C2H2-ZF gene family in the poly- and mono-embryonic citrus species and validation of the positive role of CsZFP7 in sporophytic apomixis. The C2H2 zinc finger (C2H2-ZF) gene family is involved in plant vegetative and reproductive development. Although a large number of C2H2 zinc-finger proteins (C2H2-ZFPs) have been well characterized in some horticultural plants, little is known about the C2H2-ZFPs and their function in citrus. In this work, we performed a genome-wide sequence analysis and identified 97 and 101 putative C2H2-ZF gene family members in the genomes of sweet orange (C. sinensis, poly-embryonic) and pummelo (C. grandis, mono-embryonic), respectively. Phylogenetic analysis categorized citrus C2H2-ZF gene family into four clades, and their possible functions were inferred. According to the numerous regulatory elements on promoter, citrus C2H2-ZFPs can be divided into five different regulatory function types that indicate functional differentiation. RNA-seq data revealed 20 differentially expressed C2H2-ZF genes between poly-embryonic and mono-embryonic ovules at two stages of citrus nucellar embryogenesis, among them CsZFP52 specifically expressed in mono-embryonic pummelo ovules, while CsZFP7, 37, 44, 45, 67 and 68 specifically expressed in poly-embryonic sweet orange ovules. RT-qPCR further validated that CsZFP7 specifically expressed at higher levels in poly-embryonic ovules, and down-regulation of CsZFP7 in the poly-embryonic mini citrus (Fortunella hindsii) increased rate of mono-embryonic seeds compared with the wild type, indicating the regulatory potential of CsZFP7 in nucellar embryogenesis of citrus. This work provided a comprehensive analysis of C2H2-ZF gene family in citrus, including genome organization and gene structure, phylogenetic relationships, gene duplications, possible cis-elements on promoter regions and expression profiles, especially in the poly- and mono-embryogenic ovules, and suggested that CsZFP7 is involved in nucellar embryogenesis.

Germplasm collections and morphological studies of Andropogon gayanus–Andropogon tectorum complex in southwestern Nigeria

Morphological studies were carried out on collections of the Andropogon gayanus–Andropogon tectorum complex from Southwestern Nigeria to provide full characterization of accessions of the two species and elucidate their population dynamics. Morphological data from selected accessions of A. gayanus and A. tectorum from different parts of Southwestern Nigeria were collected and characterized using an adaptation of the Descriptors for Wild and Cultivated Rice (Oryza spp.), Biodiversity International. A preliminary morphological description of the accessions was carried out at the point of collection. Garden populations were raised from the vegetative parts and maintained in Botanical Garden of Obafemi Awolowo University, Ile- Ife. The data obtained were subjected to analysis of variance and Principal Component Analysis was carried out to assess relationships among traits of the Andropogon gayanus and Andropogon tectorum. The results revealed a distinct distribution pattern of the two species of Andropogon in the area of study which suggests a south-ward migration of Andropogon gayanus; from the northern vegetational zones of Nigeria to the southern ecological zones. The migration of A. gayanus around Igbeti with occasional occurences of A. tectorum along the roadsides without any distinct phenotypic hybrid, and Budo-Ode in Oyo State is established as the southern limit of the spread of A. gayanus. This migration of A. gayanus to the south is not an invasion but a slow process. There was no A. gayanus encountered in Osun, Ondo, Ekiti and Ogun States. The plants can spread by means of propagules even if they do not produce sexual or apomictic seeds. This potential for vegetative propagation in addition to the perennial habit confers considerable advantage for colonization by the Andropogon gayanus–Andropogon tectorum complex.

Dispersal and establishment traits provide a colonization advantage for a polyploid apomictic plant.

Apomictic plants (reproducing asexually through seed) often have larger ranges and occur at higher latitudes than closely related sexuals, a pattern known as geographical parthenogenesis (GP). Explanations for GP include differences in colonizing ability due to reproductive assurance and direct/indirect effects of polyploidy (most apomicts are polyploid) on ecological tolerances. While life history traits associated with dispersal and establishment also contribute to the potential for range expansion, few studies compare these traits in related apomicts and sexuals. We investigated differences in early life history traits between diploid-sexual and polyploid-apomictic Townsendia hookeri (Asteraceae), which displays a classic pattern of GP. Using lab and greenhouse experiments, we measured seed dispersal traits, germination success, and seedling size and survival in sexual and apomictic populations from across the range of the species. While theory predicts that trade-offs between dispersal and establishment traits should be common, this was largely not the case in T. hookeri. Apomictic seeds had both lower terminal velocity (staying aloft longer when dropped) and higher germination success than sexual seeds. While there were no differences in seedling size between reproductive types, apomicts did, however, have slightly lower seedling survival than sexuals. These differences in early life history traits, combined with reproductive assurance conferred by apomixis, suggest that apomicts achieve a greater range through advantages in their ability to both spread and establish.

Human zygotic genome activation is initiated from paternal genome

Although parental genomes undergo extensive epigenetic reprogramming to be equalized after fertilization, whether they play different roles in human zygotic genome activation (ZGA) remains unknown. Here, we mapped parental transcriptomes by using human parthenogenetic (PG) and androgenetic (AG) embryos during ZGA. Our data show that human ZGA is launched at the 8-cell stage in AG and bi-parental embryos, but at the morula stage in PG embryos. In contrast, mouse ZGA occurs at the same stage in PG and AG embryos. Mechanistically, primate-specific ZNF675 with AG-specific expression plays a role in human ZGA initiated from paternal genome at the 8-cell stage. AG-specifically expressed LSM1 is also critical for human maternal RNA degradation (MRD) and ZGA. The allelic expressions of ZNF675 and LSM1 are associated with their allelically epigenetic states. Notably, the paternally specific expressions of ZNF675 and LSM1 are also observed in diploid embryos. Collectively, human ZGA is initiated from paternal genome.

Selection criteria and index analysis for the S2 maize lines of double-crosses

Abstract. Akfindarwan AK, Farid M, Syaiful SA, Anshori MF, Nur A. 2023. Selection criteria and index analysis for the S2 maize lines of double-crosses. Biodiversitas 24: 192-199. This study aimed to determine the selection criteria and the selection index for the S2 maize lineage from double cross crosses and to select the potential lines that could be continued in the S3 generation. The research was carried out in the field from July to November 2020 at the Experimental Farm of the Agriculture Faculty, Hasanuddin University, Tamalanrea District, Makassar City, South Sulawesi. The maize lines used were derived from 4 double cross populations of S1 generation, namely: DC1 (P36/B9//NK7328/JH2) with 19 genotypes, DC2 (B9)/HJ28/NK728), DC3 (B18/B2/B2/N29), and DC4 (JH45/B19/N29/B2). The results of this study revealed that the selection criteria suitable for this population were the weight of seeds per cob, peel cob weight, and weight of 100 seeds. Furthermore, the selection index analysis showed that there were 61 promising lines for F2 double cross maize that had a positive index value, while based on the Z value, there were 44 lines that had a better index value than the best parents. Therefore, it is recommended that the 61 S2 Maize lines from double crosses be further tested on to the S3 generation.

Reproduction Modes and Conservation Implications in Three Polyploid Sorbus Stenoendemics in Eastern Slovakia (Central Europe).

The remarkable species diversity of the genus Sorbus is a result of polyploidization and frequent hybridization between interacting species of different cytotypes. Moreover, hybridization is possible between several parental taxa. Gametophytic apomixis, which is common among polyploid Sorbus taxa, indicates the role of clonal reproduction in the evolutionary stabilization of hybridogeneous genotypes. The precise determination of the origin of seeds and their quantitative evaluation may elucidate inter-cytotype interactions, the potential role of mixed-cytotype populations in evolutionary success, and the long-term survival of some hybrid species. We investigated the reproduction modes of selected species of Sorbus in mixed-cytotype populations in eastern Slovakia, Central Europe. We determined the pollen quality, seed production rate, and the ploidy level of mature trees, as well as the origin of the embryo and endosperm in seeds of the stenoendemics S. amici-petri, S. dolomiticola, and S. hornadensis. The tetraploids S. amici-petri and S. hornadensis are characterized by regular and highly stainable pollen grains and reproduce predominantly via pseudogamous apomixis. In contrast, triploid S. dolomiticola usually has oval, heterogenous, and weakly stainable pollen grains, suggesting male meiotic irregularities. Although seeds originate via pseudogamous apomixis in S. dolomiticola as well, the ploidy level of sperm cells participating in the fertilization of central cells is usually determined by co-occurring species of different cytotypes. This suggests that maintaining mating partners is necessary for the long-term survival of a triploid species. We documented rare BIII hybrids and the residual sexuality in tetraploids. The distribution of seeds of meiotic and apomeiotic origins in S. amici-petri shows bimodal characteristics; however, genotypes with predominantly sexual seed types are rare. Reproduction modes documented in polyploid stenoendemics of Sorbus and inferred microevolutionary intercytotype relationships highlight the mixed-cytotype populations as the source of biodiversity in apomictic plant complexes. We suggest that conservation efforts should focus on maintaining the species and cytotypic diversity of Sorbus populations, especially when it comes to the conservation of triploid species.

Simultaneous Stenting With Edwards SAPIEN Transcatheter Pulmonary Valve Replacement.

Prestenting of the landing zone for transcatheter pulmonary valve replacement (TPVR) with a balloon-expandable valve can dilate a stenotic right ventricular outflow tract (RVOT), prevent paravalvar leak (PVL), and protect against conduit tear. Simultaneous stenting (SS) with the Melody valve has been described, but to our knowledge, SS with a SAPIEN valve has not been reported. We report our experience with this novel technique. A retrospective chart review of patients who underwent TPVR at Rady Children's hospital and UCSD Medical Center was performed. Patients were included if they had underwent SAPIEN TPVR with SS. Rationale for stent choice was a bare metal stent to relieve long-segment stenosis and covered stents to prevent PVL or to protect against conduit tear. A total of 17 cases were identified. The majority of RVOTs were transannular patches (n = 9, 56%), with a minimum diameter of 19.6 ± 5.2 mm, and the most common valve placed was an Edwards SAPIEN 26.0 mm (n = 10, 59%). All SAPIEN valves placed were of the S3 generation. The procedure was successful in all patients, with no conduit tears. Minor complications occurred in 3 patients (17.6%). Simultaneous stent deployment with a SAPIEN TPVR is an alternative 1-step technique for patients who require prestenting. SS simplifies the procedure, has low complication rates, and offers the benefits of a longer landing zone and decreased PVL.

Autoploid origin and rapid diploidization of the tetraploid Thinopyrum elongatum revealed by genome differentiation and chromosome pairing in meiosis.

Polyploidy is a common mode of evolution in flowering plants. Both the natural tetraploid Thinopyrum elongatum and the diploid one from the same population show a diploid-like pairing in meiosis. However, debate on the chromosome composition and origin of the tetraploid Th. elongatum is ongoing. In the present study, we obtained the induced tetraploid Th. elongatum and found that the induced and natural tetraploids are morphologically close, except for slower development and lower seed setting. Using probes developed from single chromosome microdissection and a Fosmid library, obvious differentiations were discovered between two chromosome sets (E1 and E2 ) of the natural tetraploid Th. elongatum but not the induced one. Interestingly, hybrid F1 derived from the two different wheat-tetraploid Th. elongatum amphiploids 8802 and 8803 produced seeds well. More importantly, analysis of meiosis in F2 individuals revealed that chromosomes from E1 and E2 could pair well on the durum wheat background with the presence of Ph1. No chromosome set differentiation on the FISH level was discovered from the S1 to S4 generations in the induced one. In metaphase of the meiosis first division in the natural tetraploid, more pairings were bivalents and fewer quadrivalents with ratio of 13.94 II + 0.03 IV (n=31). Chromosome pairing configuration in the induced tetraploid is 13.05 II + 0.47 IV (n=19), with the quadrivalent ratio being only slightly higher than the ratio in the natural tetraploid. Therefore, the natural tetraploid Th. elongatum is of autoploid origin and the induced tetraploid Th. elongatum evolutionarily underwent rapid diploidization in the low generation.

Establishment of Transcriptional Gene Silencing Targeting the Promoter Regions of GFP, PDS, and PSY Genes in Cotton using Virus-Induced Gene Silencing.

Virus-induced gene silencing (VIGS) by deploying viral-based vectors such as tobacco rattle virus (TRV) is a homology-based gene silencing technique in post-transcriptional gene silencing (PTGS) and transcriptional gene silencing (TGS) to validate the function of particular genes. The study presented here showed the induction of DNA methylation in the promoter regions of three phenotypic marker genes in different cotton accessions, including two endogenous genes such as phytoene desaturase (PDS) and phytoene synthase (PSY), and an exogenous gene, such as green fluorescent protein (GFP). First, DNA methylation was established in transgenic GFP cotton where methylation persisted up to S3 generation. Afterward, the promoter of PSY was targeted following the same conditions. Significant silencing of PSY was observed and methylation of the promoter was found up to S2 generation in red leaf cotton as detected in GFP cotton. Silencing of PDS resulted in a photobleaching phenotype; interestingly, the strength of this phenotype was diverse within the plants and was not observed in the next generation. Bisulfite sequencing results showed methylation percentage of the cytosine residues was high at CG and CHG sites of the targeted promoter sequences in the silenced plants. The findings of this paper suggest that TRV-based vector system can be used to monitor DNA methylation for both exogenous and endogenous gene levels in cotton and offer a very useful tool for plant epigenetic modification.

Reproductive Biology Factors Hampering Lemon [Citrus limon (L.) Burm. f.] Genetic Improvement

Background: Floral sterility and nucellar embryony are peculiar traits of several Citrus species and represent an obstacle to traditional breeding. Morphological sterility mainly results in pistil abortion and anther atrophy, while polyembryony is due to a mechanism known as sporophytic apomixis, which consists of the presence of embryos in the seed generated from the nucellar (maternal) tissue alongside the zygotic embryo (sexual origin). Considering the growing interest in lemon [Citrus limon (L.) Burm. f.] breeding, and the lack of information on floral sterility and the polyembryony trait among different lemon cultivars, a morphological and molecular characterization of these traits of interest was performed on forty Sicilian and international lemon cultivars available in the citrus germplasm collection of Catania University (Italy). Methods: Eight traits related to the reproductive biology were assessed on the selected lemon cultivars, namely: pistil abortion and anther atrophy, number of seeds per fruit, number of embryos per seed, percentage of seeds showing polyembryony, germination, percentage of seeds resulting in more than one plantlet, and average seed weight. Moreover, seedlings recovered after the germination assay were genotyped with SNP and SSR markers for ascertaining their nucellar or zygotic origin. In addition, PCR analysis were performed to assess the allele combination of the miniature inverted-repeat transposable element (MITE) insertion in CitRKD1, a gene associated with the occurrence of apomixis in citrus. Results: All traits showed high variability among the accessions analyzed. As for polyembryony, lemon ‘Adamopoulos’ scored the highest percentage of polyembryonic seeds (67.6%), whilst lemon ‘Lunario’ showed the lowest value (8.7%). Conclusions: Insights on the level of polyembryony within lemon varieties will represent a valuable tool for breeders for the set-up of novel mating schemes. In fact, when a polyembryonic female parent is used in cross breeding, the selection of the zygotic individual is hampered by the presence of a nucellar one.

Seed size, endosperm and germination variation in sexual and apomictic Boechera.

Asexual reproduction results in offspring that are genetically identical to the mother. Among apomictic plants (reproducing asexually through seeds) many require paternal genetic contribution for proper endosperm development (pseudogamous endosperm). We examined phenotypic diversity in seed traits using a diverse panel of sexual and apomictic accessions from the genus Boechera. While genetic uniformity resulting from asexual reproduction is expected to reduce phenotypic diversity in seeds produced by apomictic individuals, pseudogamous endosperm, variable endosperm ploidy, and the deviations from 2:1 maternal:paternal genome ratio in endosperm can all contribute to increased phenotypic diversity among apomictic offspring. We characterized seed size variation in 64 diploid sexual and apomictic (diploid and triploid) Boechera lineages. In order to find out whether individual seed size was related to endosperm ploidy we performed individual seed measurements (projected area and mass) using the phenoSeeder robot system and flow cytometric seed screen. In order to test whether individual seed size had an effect on resulting fitness we performed a controlled growth experiment and recorded seedling life history traits (germination success, germination timing, and root growth rate). Seeds with triploid embryos were 33% larger than those with diploid embryos, but no average size difference was found between sexual and apomictic groups. We identified a maternal effect whereby chloroplast lineage 2 had 30% larger seeds than lineage 3, despite having broad and mostly overlapping geographic ranges. Apomictic seeds were not more uniform in size than sexual seeds, despite genetic uniformity of the maternal gametophyte in the former. Among specific embryo/endosperm ploidy combinations, seeds with tetraploid (automomous) endosperm were on average smaller, and the proportion of such seeds was highest in apomicts. Larger seeds germinated more quickly than small seeds, and lead to higher rates of root growth in young seedlings. Seed mass is under balancing selection in Boechera, and it is an important predictor of several traits, including germination probability and timing, root growth rates, and developmental abnormalities in apomictic accessions.

Comparative Analysis of Pedigree-Based BLUP and Phenotypic Mass Selection for Developing Elite Inbred Lines, Based on Field and Simulated Data

Pedigree-based best linear unbiased prediction (pBLUP) is an effective method for genetic evaluation. The objective of this study was to assess the efficacy of pBLUP to develop superior inbred lines, using field and simulated data. This study was based on the data of 30,986 plants from six S0 generations and eight inbred progeny trials (S1–S4), performed over eight years. We measured the grain yield and the expansion volume (EV). We also simulated the EV and grain yield of 5000 plants (S0–S4) in two in silico populations, assuming 300 and 400 genes distributed across 10 chromosomes of 200 cM, respectively, and 10% of pleiotropic genes. We selected selfed plants based on the predicted additive value for EV by fitting the individual model in inbred population. The use of pBLUP provided total genetic gains in EV in the range of 1 to 45%, inversely proportional to the level of improvement of the population, and indirect changes in grain yield in the range of −17 to 3%. Only from the analysis of the in silico populations, assuming selection based on the true additive value and genetic gain computed from the genotypic values, there was evidence that pBLUP is superior to phenotypic mass selection.