Profiling Cultivars Development in Kersting's Groundnut [Macrotyloma geocarpum (Harms) Maréchal and Baudet] for Improved Yield, Higher Nutrient Content, and Adaptation to Current and Future Climates

Kersting's groundnut [Macrotyloma geocarpum (Harms.) Maréchal and Baudet], Fabaceae, is an important source of protein and essential amino acids. As a grain legume species, it also contributes to improving soil fertility through symbiotic nitrogen fixation. However, the crop is characterized by a relatively low yield (≤500 kg/ha), and limited progress has been made so far, toward the development of high-yielding cultivars that can enhance and sustain its productivity. Recently, there was an increased interest in alleviating the burdens related to Kersting's groundnut (KG) cultivation through the development of improved varieties. Preliminary investigations assembled germplasms from various producing countries. In-depth ethnobotanical studies and insightful investigation on the reproductive biology of the species were undertaken alongside morphological, biochemical, and molecular characterizations. Those studies revealed a narrow genetic base for KG. In addition, the self-pollinating nature of its flowers prevents cross-hybridization and represents a major barrier limiting the broadening of the genetic basis. Therefore, the development of a research pipeline to address the bottlenecks specific to KG is a prerequisite for the successful expansion of the crop. In this paper, we offer an overview of the current state of research on KG and pinpoint the knowledge gaps; we defined and discussed the main steps of breeding for KG' cultivars development; this included (i) developing an integrated genebank, inclusive germplasm, and seed system management; (ii) assessing end-users preferences and possibility for industrial exploitation of the crop; (iii) identifying biotic and abiotic stressors and the genetic control of responsive traits to those factors; (iv) overcoming the cross-pollination challenges in KG to propel the development of hybrids; (v) developing new approaches to create variability and setting adequate cultivars and breeding approaches; (vi) karyotyping and draft genome analysis to accelerate cultivars development and increase genetic gains; and (vii) evaluating the adaptability and stability of cultivars across various ecological regions.

The genetic variation and relationship among the natural hybrids of Mangifera casturi Kosterm

Mangifera casturi Kosterm., a mango plant from Kalimantan Selatan, Indonesia, has limited genetic information, severely limiting the research on its genetic variation and phylogeny. We collected M. casturi’s genomic information using next-generation sequencing, developed microsatellite markers and performed Sanger sequencing for DNA barcoding analysis. These markers were used to confirm parental origin and genetic diversity of M. casturi hybrids. The clean reads of the Kasturi accession were assembled de novo, producing 259 872 scaffolds (N50 = 1 445 bp). Fourteen polymorphic microsatellite markers were developed from 11 040 microsatellite motif-containing sequences. In total, 58 alleles were produced with a mean of 4.14 alleles per locus. Microsatellite marker analysis revealed broad genetic variation in M. casturi. Phylogenetic analysis was performed using internal transcribed spacers (ITS), matK, rbcL, and trnH-psbA. The phylogenetic tree of chloroplast markers placed Kasturi, Cuban, Pelipisan, Pinari, and Hambawang in one group, with M. indica as the female ancestor. Meanwhile, the phylogenetic tree of ITS markers indicated several Mangifera species as ancestors of M. casturi. Thus, M. casturi very likely originated from the cross-hybridization of multiple ancestors. Furthermore, crossing the F1 hybrids of M. indica and M. quadrifida with other Mangifera spp. may have generated much genetic variation. The genetic information for M. casturi will be a resource for breeding improvement, and conservation studies.

Entomopathogenic Nematodes: Their Characterization, Bio-Control Properties and New Perspectives

The insect parasitoid nematodes are a means boon to agronomy and serve as important bio-pesticides for controlling crop damaging insect pests. These nematodes inhabit moist soils and have been to exist in all the continents excluding Polar Regions. These nematodes have 3rd larval stage infective which is the only free living stage existing outside the host. These infective stages are mutually associated with bacteria which reside in their alimentary canal and duo are responsible for mortality of the insect host. These nematodes are currently given great attention by scientific community because of their insect killing properties and can be used to replace hazardous pesticides. These nematodes include various species belonging to genus Heterorhabditis and Steinernema, and members of insectivorous group of genus Oscheius. Before their use as bio-control agents, these nematodes need to be properly identified. Currently, these nematodes are characterized by using morphological and morphometrical parameters and advanced molecular tools including cross hybridization and scanning electron microscope studies. Their associated bacterial partners are studied through advanced molecular and biochemical techniques. The properly characterized nematodes having more entomopathogenic properties can be easily mass produced through in vitro and in vivo methods. They can be formulated in various carrier materials and supplied to farmers for effective control of damaging insect pests. Several countries have formulated various useful products of entomopathogenic nematodes which are available in markets for use by the farmer community and some have given very effective results. India is still at the early stage in the use of these nematodes for bio-control of insects in agronomy. More research in this field needs to be carried, especially in India to produce effective indigenous nematode products which may prove a boon for agriculture.

Phenotypic Characterization and RT-qPCR Analysis of Flower Development in F1 Transgenics of Chrysanthemum × grandiflorum

Gene silencing is the epigenetic regulation of any gene in order to prevent gene expression at the transcription or translation levels. Among various gene silencing techniques, RNA silencing (RNAi) is notable gene regulation technique that involves sequence-specific targeting and RNA degradation. However, the effectiveness of transgene-induced RNAi in F1 generation of chrysanthemum has not been studied yet. In the current study, we used RNAi-constructed CmTFL1 (white-flowered) and CmSVP overexpressed (yellow flowered) transgenic plants of previously conducted two studies for our experiment. Cross hybridization was performed between these intergeneric transgenic and non-transgenic plants of the winter-growing chrysanthemum selection “37” (light pink flowered). The transgene CmSVP was confirmed in F1 hybrids by RT-PCR analysis, whereas hybrids of CmTFL1 parental plants were non-transgenic. Besides this, quantitative real-time PCR (qPCR) was used to explain the molecular mechanism of flower development using reference genes. Intergeneric and interspecific hybrids produced different colored flowers unlike their respective parents. These results suggest that generic traits of CmSVP overexpressed plants can be transferred into F1 generations when crossed with mutant plants. This study will aid in understanding the breeding phenomenon among intergeneric hybrids of chrysanthemum plants at an in vivo level, and such transgenics will also be more suitable for sustainable flower yield under a low-light production system.

Development of DNA Markers From Physically Mapped Loci in Aegilops comosa and Aegilops umbellulata Using Single-Gene FISH and Chromosome Sequences

Breeding of agricultural crops adapted to climate change and resistant to diseases and pests is hindered by a limited gene pool because of domestication and thousands of years of human selection. One way to increase genetic variation is chromosome-mediated gene transfer from wild relatives by cross hybridization. In the case of wheat (Triticum aestivum), the species of genus Aegilops are a particularly attractive source of new genes and alleles. However, during the evolution of the Aegilops and Triticum genera, diversification of the D-genome lineage resulted in the formation of diploid C, M, and U genomes of Aegilops. The extent of structural genome alterations, which accompanied their evolution and speciation, and the shortage of molecular tools to detect Aegilops chromatin hamper gene transfer into wheat. To investigate the chromosome structure and help develop molecular markers with a known physical position that could improve the efficiency of the selection of desired introgressions, we developed single-gene fluorescence in situ hybridization (FISH) maps for M- and U-genome progenitors, Aegilops comosa and Aegilops umbellulata, respectively. Forty-three ortholog genes were located on 47 loci in Ae. comosa and on 52 loci in Ae. umbellulata using wheat cDNA probes. The results obtained showed that M-genome chromosomes preserved collinearity with those of wheat, excluding 2 and 6M containing an intrachromosomal rearrangement and paracentric inversion of 6ML, respectively. While Ae. umbellulata chromosomes 1, 3, and 5U maintained collinearity with wheat, structural reorganizations in 2, 4, 6, and 7U suggested a similarity with the C genome of Aegilops markgrafii. To develop molecular markers with exact physical positions on chromosomes of Aegilops, the single-gene FISH data were validated in silico using DNA sequence assemblies from flow-sorted M- and U-genome chromosomes. The sequence similarity search of cDNA sequences confirmed 44 out of the 47 single-gene loci in Ae. comosa and 40 of the 52 map positions in Ae. umbellulata. Polymorphic regions, thus, identified enabled the development of molecular markers, which were PCR validated using wheat-Aegilops disomic chromosome addition lines. The single-gene FISH-based approach allowed the development of PCR markers specific for cytogenetically mapped positions on Aegilops chromosomes, substituting as yet unavailable segregating map. The new knowledge and resources will support the efforts for the introgression of Aegilops genes into wheat and their cloning.

Robust Response to Plum pox virus Infection via Plant Biotechnology

Our goal was to target silencing of the Plum pox virus coat protein (PPV CP) gene independently expressed in plants. Clone C-2 is a transgenic plum expressing CP. We introduced and verified, in planta, the effects of the inverse repeat of CP sequence split by a hairpin (IRSH) that was characterized in the HoneySweet plum. The IRSH construct was driven by two CaMV35S promoter sequences flanking the CP sequence and had been introduced into C1738 plum. To determine if this structure was enough to induce silencing, cross-hybridization was made with the C1738 clone and the CP expressing but PPV-susceptible C2 clone. In total, 4 out of 63 clones were silenced. While introduction of the IRSH is reduced due to the heterozygous character in C1738 plum, the silencing induced by the IRSH PPV CP is robust. Extensive studies, in greenhouse containment, demonstrated that the genetic resource of C1738 clone can silence the CP production. In addition, these were verified through the virus transgene pyramiding in the BO70146 BlueByrd cv. plum that successfully produced resistant BlueByrd BO70146 × C1738 (HybC1738) hybrid plums.

Spikelet movements, anther extrusion and pollen production in wheat cultivars with contrasting tendencies to cleistogamy

Background Cleistogamic flowers are a main barrier in pollen dispersal for cross-pollination necessary in wheat hybrid breeding. The aim of our study was to gain new knowledge on the biology of wheat flowering, in particular on the differences between the cleisto- and chasmogamic forms which has certainly cognitive significance, but it can also be used in practice when seeking a female and male ideotypes for cross hybridization. Results We characterized the most significant features defining the flowering specificity in two wheat cultivars with contrasting tendency to cleistogamy: Piko (chasmogamous) and Dacanto (cleistogamous). In the field observations we assessed diurnal pattern of anther extrusion and anther extrusion capacity. For the first time we adapted the time lapse method for measuring kinetics of the spikelet movement and 3-D image correlation technique for the non-invasive measurements of potential deformations of the spikelet lemmas. We found that the two cultivars differ in the potential of pollen dispersion for-cross-pollination and in the spikelet kinetics. We also described some anatomical traits that can have potential functional role in floret opening. None of the cultivars showed any symptoms of lemma surface deformation. Conclusions The cleistogamic and chasmogamic wheat cultivars differ significantly in the potential for pollen dispersion for cross-pollination, which is mainly related to anther extrusion capacity. Although none of these features differentiated the cultivars clearly, we assume, based on spikelet kinetics and the lack of lemmas surface deformation, that the water transport and turgor of cells is essential for the floret opening and anther extrusion in wheat. The search for parental ideotype should be supported by marker assisted selection, e.g. based of polymorphisms in genes related to aquaporin biosynthesis.

Genomic Data Reveal the Genetic Variation among Natural Mangifera Casturi Kosterm. Hybrids, an Underutilized Fruit Tree Under ‘Extinct in Wild’ Status from Kalimantan Selatan, Indonesia

Background: Mangifera casturi Kosterm. is an endemic local mango fruit from Kalimantan Selatan, Indonesia. The limited genetic information available on this fruit has severely limited the scope of research into its genetic variation and phylogeny. This study aimed to collect genomic information from M. casturi using next-generation sequencing technology and to develop microsatellite markers and perform Sanger sequencing for DNA barcoding analysis.Results: The clean reads of the Kasturi accession of M. casturi were assembled de novo using a Ray assembler, producing 259,872 scaffolds with an N50 value of 1,445 bp. Fourteen polymorphic microsatellite markers were developed from 11,040 sequences containing microsatellite motifs. In total, 55 alleles were produced, and the mean number of alleles per locus was 3.93. Results from the microsatellite marker analysis revealed broad genetic variation in M. casturi. Phylogenetic analysis was performed using internal transcribed spacers (ITS), matK, rbcL, and trnH-psbA. The phylogenetic tree of chloroplast markers showed that Kasturi, Mawar, Pelipisan, Pinari, and Hambawang belong to one group, with M. indica as the female ancestor. In comparison, the phylogenetic tree of ITS markers indicated several Mangifera species as multiple ancestors of M. casturi. Conclusions: This study strongly suggested that M. casturi originated from the cross-hybridization of multiple ancestors. Further, crossing the F1 hybrids of M. indica and M. quadrifida with other Mangifera spp. was hypothesized to produce the observed high genetic variation. The genetic information for this fruit is also a resource for the breeding and improvement as well as for conservation studies of this species.

Variability and inheritance of fiber length and wilt resistance in a complex 4-5 specific and backcross hybridization of cotton

In this paper presents data of researches about fiber length in the 4-5 species. The largest number of plants with the indicated analyzed trait in hybrid combinations, obtained with the participation of the variety Omad, appeared with a number of variations in fiber length at 35,0-36,0 mm and above. Investigation of 4-species hybrids of F2 and back cross hybridization allowed some improvement in the average value of trait when compared with F1. Despite the emergence of transgressive plants with fiber length 36,1-37,0 mm and above, the average trait of composite 5 species hybrids F2 remained almost at the level of F1, i.e. 35.5-36.0 mm. The greatest number of positive recombinants occurred in 5 species back crossing developed with the participation of Termez-31.

Variability and inheritance of fiber length and wilt resistance in a complex 4-5 specific and backcross hybridization of cotton

In this paper presents data of researches about fiber length in the 4-5 species. The largest number of plants with the indicated analyzed trait in hybrid combinations, obtained with the participation of the variety Omad, appeared with a number of variations in fiber length at 35,0-36,0 mm and above. Investigation of 4-species hybrids of F2 and back cross hybridization allowed some improvement in the average value of trait when compared with F1. Despite the emergence of transgressive plants with fiber length 36,1-37,0 mm and above, the average trait of composite 5 species hybrids F2 remained almost at the level of F1, i.e. 35.5-36.0 mm. The greatest number of positive recombinants occurred in 5 species back crossing developed with the participation of Termez-31.