Elite Germplasm Research Articles

Prospects of Developing Novel Genetic Resources by Chemical and Physical Mutagenesis to Enlarge the Genetic Window in Bread Wheat Varieties

Sustainable production and improved genetic gains can be achieved by broadening the genetic window of elite wheat germplasm. Here, we induced mutations in two spring wheat varieties, viz., NN-Gandum-1 (NN-1) and Punjab-11 (Pb-11), by exposing their seeds to ethyl methane sulfonate (EMS) and γ-rays, respectively. We characterized >3500 lines of each NN-1 and Pb-11 derived population in three consecutive generations, viz., M5, M6, and M7, for important traits, e.g., plant height, heading date, spike morphology and rust resistance. We observed significant genetic variation and correlations in both populations for all investigated traits. We observed differences in terms of number of mutants between NN-1 (22.76%) and Pb-11 (26.18%) which could be ascribed to the genotype-by-mutagen interaction. High broad-sense heritability (H2) estimates, that are vital for higher genetic gains, were observed for all of the investigated traits in both populations (H2 = 0.69–0.91 in NN-1 and 0.84–0.98 in Pb-11). Particularly, to breed for rust resistance, we selected a subset (n = 239) of M7 lines that also showed phenotypic variation for other traits. Our studies (1) show the relevance to artificial mutagenesis to create genetic variation in elite germplasm for their immediate use in current breeding programs, and (2) provide material for downstream identification of genes associated with traits of high agronomic importance.

Editorial: Strengthening the researches of genomics of bast fiber crops to promote elite allele mining and germplasm innovation

Editorial: Strengthening the researches of genomics of bast fiber crops to promote elite allele mining and germplasm innovation

Association mapping of sponge cake volume in U.S. Pacific Northwest elite soft white wheat (Triticum aestivum L.)

Sponge cake quality is an essential end-use trait for U.S. Pacific Northwest (PNW) soft white (SW) wheat, primarily measured by the volume of a cake. The baking of each cake is relatively expensive, labor intensive, and requires a substantial amount of flour. As a result, sponge cake baking is conducted in later generations on a limited number of breeding lines. Insight into the genetic architecture of sponge cake volume would help provide an alternative to cake baking. Here we describe the analysis of 387 wheat genotypes spanning 27 crop years with a total of 685 environments for sponge cake volume. Genotyping by sequencing was used to generate molecular markers in wheat representing elite germplasm of the PNW. A genome wide association study was conducted to discover marker trait associations, while accounting for population structure and kinship. A total of six significant marker trait associations were identified on chromosomes 1AB, 2A, 3BD, 4A, and 6A. Collectively these marker trait associations explained 29.9% (ranging from 0.3 to 14.2%) of the sponge cake volume variation. Previous research detected associations with agronomic, grain, flour, milling and baking traits on each of these chromosomes, however, none for sponge cake volume. This study serves as a foundation experiment by providing insights into the genetic architecture of sponge cake volume in a historical dataset.

Trait Based Modelling Approach for Selection of Elite Germplasm Accessions in Soybean [Glycine max (L). Merrill

Background: Identification of suitable factors that influence significantly to the response is crucial for the traits based breeding program to make a better decision about improvement in productivity. Multiple linear regression (MLR) is the benchmark method commonly using to identify suitable factors for crop improvement. It doesn’t work always due to stringent assumption (Multicollinearity, Linearity) behind the MLR model. Here we tried to develop an efficient model for the selection of major traits that contribute to seed yield in soybean by comparing different models.Methods: Field experiment was conducted using 98 soybean core population through augmented design.18 morphometric traits obtain from soybean core population were considered under the study as regressors.Multiple linear regression (MLR), Principle component Regression (PCR), Regression tree and Random Forest models were compared to select traits based on prediction accuracy.Result: All the models identified the number of pods per plant (NPP) has the most influencing variable to the soybean yield. However random forest has a much higher prediction power (RMSE=4.59, MAPE=0.18) compared to other models under study. The results of random forest revealed that the number of pods per plant, number of branches per plant and other associated characters like plant height at harvest as highly influencing traits for seed yield in soybean.Finally, tried to identify genotypesthat possess superiority about most influencing morphological characters on seed yield using cluster analysis.

Genetic diversity of Indian garlic core germplasm using agro-biochemical traits and SRAP markers.

The characterization of garlic germplasm improves its utility, despite the fact that garlic hasn't been used much in the past. Garlic has an untapped genetic pool of immense economic and medicinal value in India. Hence, using heuristic core collection approach, a core set of 46 accessions were selected from 625 Indian garlic accessions based on 13 quantitative and five qualitative traits. The statistical measures (CV per cent, CR per cent, VR per cent) were used to sort the core set using Shannon-Wiener diversity index and the Nei diversity index. In addition, the variation within the core set was tested for 18 agro-morphological and six biochemical characteristics (allicin, phenol content, pyruvic acid, protein, allyl methyl thiosulfinate (AMTHS), and methyl allyl thiosulfinate (MATHS)). Further study of the core set's molecular diversity was performed using sequence related amplified polymorphism (SRAP) markers, which revealed a wide range of diversity among the core set's accessions, with an average polymorphism efficiency (PE) of 80.59 percent, polymorphism information content (PIC) of 0.29, effective multiplex ratio (EMR) of 3.51, and marker index (MI) of 0.99. The findings of this study will be useful in identifying high-yielding, elite garlic germplasm lines with the trait of interest. Since this core set is indicative of total germplasm, these selected breeding lines will be used for genetic improvement of garlic in the future.

Influence of rootstocks on scion leaf mineral content in mango tree (Mangifera indica L.)

Grafting is an important practice in mango (Mangifera indica L.) orchard management for propagating elite germplasm but there is limited information on the effect of rootstock-scion interactions on the mineral nutrition of mango. Here, we analyzed the variations in leaf nutrients of commercial (‘B74’ and ‘Kensington Pride’ [KP]) and newly developed cultivars from the National Mango Breeding Program (NMBP) (‘NMBP1243’, ‘NMBP1201’, and ‘NMBP4069’) grafted onto six different rootstocks (‘B’, ‘KP’, ‘Mallapunyah’, ‘SabahBRF’, ‘Vellaikulamban’, and ‘Watertank’) to determine the influence of the rootstock on the scion leaf nutrient content. Deviation from the optimum percentage (DOP and ΣDOP indexes) was calculated to compare nutrition status/balance among the different rootstocks. Leaf nutrient concentrations and their DOP index were affected by the rootstocks and scions. None of the rootstocks or scions had consistently higher or lower nutrient levels, and interaction effects between rootstock and scion factors were insignificant. The DOP index indicated an imbalance of nutrients in the leaves, but scions grafted onto rootstocks ‘Vellaikulamban’ and ‘SabahBRF’ were more nutritionally balanced, indicating that they are preferred rootstocks for propagation. Principal component analysis showed a clear separation among rootstocks according to their differences in nutrient concentrations, such that ‘Vellaikulamban’ and ‘SabahBRF’ (Group 1) were different from ‘Watertank’ and ‘B’ (Group 2) as well as from ‘KP’ and ‘Mallapunyah’ (Group 3). Scions were clustered based on their similarities with ‘B74’ and ‘NMBP1201’ in Cluster 1, ‘NMBP 4069’ in Cluster 2, ‘NMBP 1243’ in Cluster 3, and ‘KP’ in Cluster 4. This study provides valuable information to understand the potential impact of the rootstock on the nutritional status of scions as well as for the selection of rootstock-scion combinations for variety improvement.

Influence of Soil Variation on Diosgenin Content Profile in Costus speciosus from Indo-Gangetic Plains.

Costus speciosus is a rich source of commercially important compound Diosgenin, distributed in different regions of India. The present investigation was aimed to quantify diosgenin through High Performance Thin Layer Chromatography in 34 germplasms of Costus speciosus and also to identify the superior sources and to correlate the macronutrients of rhizospheric soil. The starch content varied in microscopic examination and correlated inversely (r=-0.266) with diosgenin content. Findings revealed that the extraction process with acid hydrolysis yielded higher diosgenin content (0.15-1.88 %) as compared to non-hydrolysis (0.009-0.368 %) procedure. Germplasms from Uttar Pradesh (NBCS-4), Jharkhand (NBCS-39) and Bihar (NBCS-2) were identified as elite chemotypes based on hierarchical clustering analysis. The phosphorous content of respective rhizospheric soil correlated positively (r=0.742) with diosgenin content. Findings of present study are useful to identify the new agrotechniques. The elite germplasms can also be used as quality planting material for large scale cultivation in order to assure a sustained supply to the herbal drug industry.

An integrative analysis of yield stability for a GWAS in a small soybean breeding population

AbstractDrought stress is one of the most important factors limiting soybean [Glycine max (L.) Merr.] productivity and reducing yield stability. Soybean breeders need phenotypic and genotypic tools to improve drought stress tolerance, but most of available strategies are expensive and unaffordable for small‐scale public breeding programs. In this study, elite germplasm of a locally adapted breeding population was used to estimate a yield stability index as an indicator of drought response. In order to associate yield stability of analyzed genotypes to drought response, water deficit scenarios related to the crop cycle group were defined. Four groups of genotypes were identified in relation to yield stability: two groups showed stables yield (without interaction with water deficit scenarios), and two groups showed unstable yield (with crossover interaction with water deficit scenarios). This phenotypic information was used to identify genomic regions and candidate genes associated with yield stability index. A new method for the definition of a quantitative trait loci (QTL) region was developed based on the probability of marker pairwise of belonging to four linkage disequilibrium (LD) categories. Seven QTL were found and their implication on drought tolerance was further supported by linkage to previously reported QTL for water use efficiency trait.

Developmental patterns and rates of organogenesis across modern and well-adapted wheat cultivars

Genetic gains in wheat yield are required to meet future demand. In order to design strategic crosses, it is essential to quantify the genetic variation among elite cultivars on traits responsible for the adaptation and the generation of yield components. We explored the degree of variation within well-adapted commercial cultivars on developmental phases (vegetative, early reproductive and late reproductive), final leaf number and phyllochron, leaf and spikelet plastochron, number of spikelets, florets, and dynamics of floret primordia development. Variation in flowering time among cultivars was minor but they differed in the partitioning of that time into particular phases. Main differences were found for the length of the late reproductive phase, partly linked to those in final leaf number. Variations on number of fertile florets at anthesis were mostly explained by floret survival, due to differential ability of labile florets to maintain normal development between cultivars. These differences were related to the duration of the floret development, but not to the synchrony of development among florets. These findings provide useful information for breeders when selecting genotypes for strategic crosses, as considering floret survival through duration of floret development in one of the parents (for which these processes shall be characterised in the elite germplasm) might increase the likelihood of transgressive segregation towards high-yielding offspring.

QTL Mapping of Cucurbit yellow stunting disorder virus Resistance in Melon Accession PI 313970

Cucurbit yellow stunting disorder virus (CYSDV) is a devastating viral disease of melon that can cause significant yield and quality losses. This disease has recently emerged as a major concern in the southwest United States and major melon-growing regions across the world. Coinfection of melon by Cucurbit chlorotic yellows virus (CCYV) was recognized in Imperial Valley and neighboring production areas of California and Arizona in 2018, but its importance remains largely unknown. Identifying and deploying CYSDV resistance from elite germplasm is an economical and effective way to manage the disease. A F2:3 population was developed from a cross of susceptible ‘Top Mark’ with CYSDV-resistant PI 313970, which was shown to possess a single recessive gene for resistance to CYSDV. The F2:3 population was phenotyped in the field in response to natural, mixed infections by the two viruses, CYSDV and CCYV in the Fall melon seasons of 2018 and 2019. Phenotypic data (foliar yellowing) from both years were not useful for mapping CYSDV resistance quantitative trait loci (QTL), as PI 313970 and CYSDV-resistant F2:3 plants exhibited yellowing symptoms from CCYV coinfection. QTL analysis of the relative titer of CYSDV calculated from reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) data identified one locus on chromosome 3 at the physical location of S5-28,571,859 bp that explained 20% of virus titer variation in 2018 but was undetected in 2019. A locus on chromosome 5 between S5-20,880,639 to S5-22,217,535 bp explained 16% and 35% of the variation in CYSDV titer in 2018 and 2019, respectively. One or both of the markers were present in six of 10 putative melon CYSDV resistance sources. Markers flanking the 2019 QTL were developed and can be used in marker-assisted breeding of CYSDV-resistant melons.

QTL Analysis of Adult Plant Resistance to Stripe Rust in a Winter Wheat Recombinant Inbred Population

Stripe rust, caused by the fungus Puccinia striiformis f. sp. tritici, is a worldwide disease of wheat that causes devastating crop losses. Resistant cultivars have been developed over the last 40 years that have significantly reduced the economic impact of the disease on growers, but in heavy infection years it is mostly controlled through the intensive application of fungicides. The Pacific Northwest of the United States has an ideal climate for stripe rust and has one of the most diverse race compositions in the country. This has resulted in many waves of epidemics that have overcome most of the resistance genes traditionally used in elite germplasm. The best way to prevent high yield losses, reduce production costs to growers, and reduce the heavy application of fungicides is to pyramid multiple stripe rust resistance genes into new cultivars. Using genotyping-by-sequencing, we identified 4662 high quality variant positions in a recombinant inbred line population of 196 individuals derived from a cross between Skiles, a highly resistant winter wheat cultivar, and Goetze, a moderately to highly susceptible winter wheat cultivar, both developed at Oregon State University. A subsequent genome wide association study identified two quantitative trait loci (QTL) on chromosomes 3B and 3D within the predicted locations of stripe rust resistance genes. Resistance QTL, when combined together, conferred high levels of stripe rust resistance above the level of Skiles in some locations, indicating that these QTL would be important additions to future breeding efforts of Pacific Northwest winter wheat cultivars.

The physiology and genetics behind fruiting efficiency: a promising spike trait to improve wheat yield potential

Fruiting efficiency (FE, grains per g of spike dry weight at anthesis) was proposed as a promising spike trait to improve wheat yield potential, based on its functional relationship with grain number determination and the evidence of trait variability in elite germplasm. During the last few years, we have witnessed great advances in the understanding of the physiological and genetic basis of this trait. The present review summarizes the recent heritability estimations and the genetic gains obtained when fruiting efficiency was measured at maturity (FEm, grains per g of chaff) and used as selection criterion. In addition, we propose spike ideotypes for contrasting fruiting efficiencies based on the fertile floret efficiency (FFE, fertile florets per g of spike dry weight at anthesis) and grain set (grains per fertile floret), together with other spike fertility-related traits. We also review novel genes and quantitative trait loci available for using marker-assisted selection for fruiting efficiency and other spike fertility traits. The possible trade-off between FE and grain weight and the genes reported to alter this relation are also considered. Finally, we discuss the benefits and future steps towards the use of fruiting efficiency as a selection criterion in breeding programs.

Ecological features and traditional knowledge of Roscoea alpina Royle a medicinal plant in Himalaya

Roscoea alpina Royle (Zingiberaceae) commonly known as Kakoli is a perennial medicinal herb endemic to the Himalayas. It is used in vitality strengthening Ayurveda groups i.e., Astavarga and Jeevaniyagana and in preparation of Ayurvedic formulations e.g., Chyavanprasha and Divya Pidantak Tail . Considering its medicinal uses, high demand and overexploitation, natural habitats were surveyed in subalpine and alpine regions of Garhwal Himalayas for population estimation and to identify elite germplasm. R. alpina frequency was recorded more than 60 % in Tungnath, Dayara, Valley of Flowers and Kedarnath populations. However, plant density and area occupied were low compared to other species of subalpine and alpine region. For threat category assessment, IUCN Red List Categories and criteria were used and conservation status was assigned, based on site to site and for entire Garhwal region of the Western Himalaya. R. alpina , based on extent of occurrence was categorized as Vulnerable and based on number of mature individuals as Endangered for the Garhwal Himalaya. Further, habitat destruction and degradation were the major threats for population reduction in the wild. Morphological variation revealed plants from Tungnath and Kedarnath may be used for future propagation and domestication efforts.

A novel micropropagation of Lycium ruthenicum and epigenetic fidelity assessment of three types of micropropagated plants in vitro and ex vitro.

Lycium ruthenicum is an excellent eco-economic shrub. Numerous researches have been conducted for the function of its fruits but scarcely focused on the somaclonal variation and DNA methylation. An efficient micropropagation protocol from leaves and stems of L. ruthenicum was developed in this study, in which not only the leaf explants but also the stem explants of L. ruthenicum were dedifferentiated and produced adventitious buds/multiple shoots on one type of medium. Notably, the efficient indirect organogenesis of stem explants was independent of exogenous auxin, which is contrary to the common conclusion that induction and proliferation of calli is dependent on exogenous auxin. We proposed that sucrose supply might be the crucial regulator of stem callus induction and proliferation of L. ruthenicum. Furthermore, results of methylation-sensitive amplified polymorphism (MSAP) showed that DNA methylation somaclonal variation (MSV) of CNG decreased but that of CG increased after acclimatization. Three types of micropropagated plants (from leaf calli, stem calli and axillary buds) were epigenetically diverged more from each other after acclimatization and the ex vitro micropropagated plants should be selected to determine the fidelity. In summary, plants micropropagated from axillary buds and leaves of L. ruthenicum was more fidelity and might be suitable for preservation and propagation of elite germplasm. Also, leaf explants should be used in transformation. Meanwhile, plants from stem calli showed the highest MSV and might be used in somaclonal variation breeding. Moreover, one MSV hotspot was found based on biological replicates. The study not only provided foundations for molecular breeding, somaclonal variation breeding, preservation and propagation of elite germplasm, but also offered clues for further revealing novel mechanisms of both stem-explant dedifferentiation and MSV of L. ruthenicum.

SSR markers assay-based establishment of distinctness, uniformity and stability of dolichos bean [Lablab purpureus (L.) Sweet var. Lignosus] advanced breeding lines and elite germplasm accessions

Registration of crop varieties under Indian PPV & FR Act requires establishment of their Distinctiveness, Uniformity and Stability (DUS). At present, DUS of crop varieties are being established using morphological traits. DNA marker assay-based DUS testing would complement that based on morphological traits. Four markers, from a total of 220 tested SSR markers were polymorphic among two advanced breeding lines and two elite germplasm accessions (used as candidate varieties), and two released varieties (used as reference varieties) of dolichos bean. The four polymorphic simple sequence repeat (SSR) markers were used to establish distinctness of candidate varieties from reference varieties based on alleles unique to candidate varieties. Uniformity of candidate varieties were established using two criteria, (1) per cent homozygous plants (> 95%) and (2) SSR-homozygous loci ratio (HLR) (> 95%).The candidates which were found uniform at tested SSR loci were deemed stable. The candidate varieties were distinct from reference varieties at two of the four tested polymorphic SSR markers, as evident by the presence of alleles unique to candidate varieties. One hundred per cent homozygous plants and SSR-HLR within all the candidate varieties at all the four tested SSR marker loci suggested uniformity and deemed-stability of candidate varieties. The study demonstrated the utility of SSR markers to establish DUS of ABL/elite germplasm accessions in dolichos bean.

Transient reprogramming of crop plants for agronomic performance.

The development of a new crop variety is a time-consuming and costly process due to the reliance of plant breeding on gene shuffling to introduce desired genes into elite germplasm, followed by backcrossing. Here, we propose alternative technology that transiently targets various regulatory circuits within a plant, leading to operator-specified alterations of agronomic traits, such as time of flowering, vernalization requirement, plant height or drought tolerance. We redesigned techniques of gene delivery, amplification and expression around RNA viral transfection methods that can be implemented on an industrial scale and with many crop plants. The process does not involve genetic modification of the plant genome and is thus limited to a single plant generation, is broadly applicable, fast, tunable and versatile, and can be used throughout much of the crop cultivation cycle. The RNA-based reprogramming may be especially useful in plant pathogen pandemics but also for commercial seed production and for rapid adaptation of orphan crops.

Genetic and biochemical stability of in vitro raised and conventionally propagated Centella asiatica - A valuable medicinal herb

An efficient micropropagation protocol was developed for C. asiatica, a commercially important herb having high medicinal and cosmetic properties. Shoot induction was observed on Murashige and Skoog (MS) medium fortified with various combinations of benzyladenine (BA) (2.22 - 22.2 µM) and naphthalene acetic acid (NAA) (1.34 - 8.06 µM). The maximum number of shoots (11.30 ± 0.68) per explant was initiated on MS media containing BA (6.66 µM) and NAA (2.69 µM). Maximum shoot proliferation was achieved on MS medium supplemented with BA (6.66 µM) and indole-3-acetic acid (IAA) (2.84 µM). The best rooting (24.69 ± 0.08) per explant was achieved on half-strength MS medium supplemented with indole-3-butyric acid (IBA) (4.92 µM). The in vitro regenerated plantlets were acclimatized and shifted to the field. Genetic stability of the micropropagated regenerants was evaluated using inter-simple sequence repeat (ISSR) markers and flow cytometry. Out of 28 primers tested, 9 primers resulted in 561 distinct and reproducible bands ranging from 200 to 1400 bp. Monomorphic banding profile was observed between the mother plant and tissue culture regenerants. The mean 2C DNA content of mother and micropropagated plants evaluated by flow cytometry were estimated to be 0.14 and 0.12 pg, thereby indicating no significant change in the 2C DNA content. Additionally, HPLC (High performance liquid chromatography) analysis did not show any significant differences in the content of triterpenes and phenolic compounds in both mother and micropropagated plants. Thus, this protocol could be implemented for large scale cultivation of elite germplasm of C. asiatica.

Morphogenic Regulators and Their Application in Improving Plant Transformation.

Generation of plant lines with transgene or edited gene variants is the desired outcome of transformation technology. Conventional DNA-based plant transformation methods are the most commonly used technology but these approaches are limited to a small number of plant species with efficient transformation systems. The ideal transformation technologies are those that allow biotechnology applications across wide genetic background, especially within elite germplasm of major crop species. This chapter will briefly review key regulatory genes involved in plant morphogenesis with a focus on in vitro somatic embryogenesis and their application in improving plant transformation.

Isolation of Staged and Viable Maize Microspores for DH Production.

Isolated microspore culture systems have been designed in maize by several groups, mainly from the late 1980s to early 2000s. However, even with optimized protocols, microspore embryogenesis induction has remained very dependent on the genotype in maize, with elite germplasm generally displaying no response or very low response. Yet, these last few years, significant progress has been accomplished in understanding and controlling microspore embryogenesis induction in model dicot and monocot species. This knowledge may be transferred to maize, and isolated microspore culture may gain new interest in this crop, at least for embryogenesis research. The methods we hereby present in detail permit the purification of 3-12×105 viable microspores per maize tassel, at the favorable stage for microspore embryogenesis. When cultured in appropriate liquid media, microspores from responsive genotypes give rise to androgenic embryos, which can then be regenerated into fertile doubled haploid plants.

Assessment of GeneticVariability and Character Association in Yield Related Traits and Yellow Vein Mosaic Virus Disease Resistance in Okra (Abelmoschus esculentus L. Moench)

Okra is India’s most widely grown tropical and subtropical vegetable crop. Despite widespread cultivation, crop yield and productivity remain subpar. Yellow vein mosaic virus (YVMV) disease is a serious threat to Indian okra production, causing yield losses ranging from 17.09 to 96.49 per cent. In this context, the present study was aimed to assess the variability and character association of 74 elite germplasm accessions for thirteen quantitative and quality traits. High phenotypic and genotypic coefficient of variation (PCV and GCV) were observed among all evaluated traits except fruit diameter, which showed a moderate PCV and GCV. All traits had high heritability and high genetic advance, implying that the heritability was most likely due to additive gene effects. Direct selection of these traits would be advantageous for okra improvement, especially in developing a YVMV resistant genotype. Fruit yield per plant exhibited a significant and positive genotypic as well as phenotypic correlation with fruit weight, fruit length, peroxidase activity, polyphenol activity, fruit diameter, total phenolic content and plant height, whereas node at which the first flower appeared and YVMV were negatively correlated to fruit yield per plant. Correlation studies indicated YVMV resistance towards high total phenolic content, polyphenol oxidase, and peroxidase. The high positive direct effects on fruit yield per plant were observed for fruit weight followed by number of fruit per plant and total phenolic content. This study revealed that simultaneous selection of these traits can help in increasing yield and YVMV resistance.