National Gene Bank Research Articles

The Draft Genome of Coelastrum proboscideum (Sphaeropleales, Chlorophyta)

Coelastrum proboscideum Bohlin, 1896 (Sphaeropleales, Scenedesmaceae, Chlorophyta) is a coenobial species with cosmopolitan distribution in diverse freshwater habitats. Coelastrum spp. are widely tested for biotechnological applications such as carotenoid and lipid production, and in bioremediation of wastewater. Here, we report the draft genome of C. proboscideum var. dilatatum strain SAG 217-2. The final assembly comprised 125,935,854 bp with over 8357 scaffolds. The whole-genome data is publicly available in the Nucleotide Sequence Archive (CNSA) of China National GeneBank (CNGB) (https://db.cngb.org/cnsa/) under the accession number CNA0014153.

Screening of 19,460 genotypes of wheat species for resistance to powdery mildew and identification of potential candidates using focused identification of germplasm strategy (FIGS)

AbstractPowdery mildew (PM) caused by Blumeria graminis (DC). E.U. Speer f.sp. tritici Em. Marchal (Syn. Erysiphe gramnis DC f.sp. tritici, Em. Marchal) is one of the important diseases of wheat worldwide. A comprehensive evaluation of wheat germplasm accessions (19,460) conserved in the National Genebank of ICAR–National Bureau of Plant Genetic Resources was conducted to identify sources of resistance to PM. Accessions belonging to the three wheat species—bread wheat (Triticum aestivum L. subsp. aestivum) (15,944), durum wheat (T. durum Desf.) (3,359), and emmer wheat (T. dicoccum Schrank ex Schübl.) (157)—were screened at Wellington, a hotspot location for PM, for two consecutive seasons. Screening results indicated that 7271 (45%) from bread wheat, 756 (22%) from durum wheat, and 22 (14%) from emmer were resistant. Out of 8094 PM‐resistant accessions, 60% were indigenous, while majority of the 40% exotic were from CIMMYT. Focused identification of germplasm strategy (FIGS), which identifies a set of similar plant genotypes with a greater possibility of containing specific target traits, was used to form a subset of 52 accessions (from 19,460) that have the potential to contain new PM resistance genes. Resistant accessions identified in the study have enriched the existing gene pool for PM resistance in wheat and will serve as a potential source for resistance in future.

Assessment of the Glutenin Subunits Diversity in a Durum Wheat (T. turgidum ssp. durum) Collection from Morocco

Landraces and old wheat cultivars display great genetic variation and constitute a valuable resource for the improvement of modern varieties, especially in terms of quality. Gluten quality is one of the major determinants of wheat quality, and it is greatly influenced by variation in the high molecular weight and low molecular weight glutenin subunits (HMW-GS and LMW-GS). Identification of novel allelic variants for either of the two groups of the gluten-forming proteins could greatly assist in the improvement of wheat gluten quality. In the present study, the allelic composition of the HMW- and LMW-GS of ninety-five durum wheat accessions was evaluated. These accessions included Moroccan cultivars and landraces and North American cultivars and were all conserved in the National Gene Bank from Morocco. In total, 20 cataloged alleles and 12 novel alleles were detected. For the HMW-GS, two alleles were found at the Glu-A1 locus, and seven different allelic variants were identified at the Glu-B1 locus. Among them, two alleles were new (alleles Glu-B1cp and co). Additionally, two of the analyzed accessions exhibited the Glu-D1d allele, suggesting the presence of the Glu-D1 locus introgression. For the LWM-GS, eight, ten and two alleles were identified at the Glu-A3, Glu-B3 and Glu-B2 loci, respectively. Among them, two new allelic variants were identified at the Glu-A3 locus, and seven new allelic variants were identified at the Glu-B3 locus. Overall, the Moroccan landraces exhibited a greater genetic diversity and a greater number of glutenin alleles compared to the Moroccan and North American durum wheat cultivars. The novel germplasm and glutenin alleles detected in this study could contribute to the improvement of durum wheat quality and the expansion of modern durum wheat genetic diversity.

Agro-morphological characterization of local garlic (Allium sativum L.) germplasm accessions collected from different regions of Jammu and Kashmir

Garlic (Allium sativum L.) is increasingly becoming an essential component in human diet due to tremendous health-promoting benefits; its traditional local types are preferred for their taste. Garlic clones have specific adaptations to different agro-climatic regions; some have adapted to specific environments through artificial and natural selection. The study was undertaken to assess the variation in some agro-morphological characters of 17 local garlic germplasm accessions collected from different regions of Jammu and Kashmir including far-flung areas of Karthaie Padder Kishtwar and Balot Bhaderwah Doda. Most of these local garlic germplasm accessions are characterized by more pungent smell and distinctive taste often with pigmented coat layers. Some of these accessions have been cryo-preserved in National Gene Bank at ICAR-NBPGR New Delhi. The results have demonstrated the existence of a good amount of agro-morphological diversity in these local garlic germplasm accessions. Different ecotypes exhibit considerable variation among themselves in agronomic characteristics. Therefore, there is a need for concerted efforts for exploration, collection and maintenance of locally available garlic germplasm possessing quality traits including disease and pest resistance besides good storability characteristics for future garlic crop improvement programmes in the region.

Droplet vitrification technique for cryopreservation of a large diversity of blackcurrant (Ribes nigrum L.) cultivars

The aim of plant gene banks is to preserve genetic resources selected based on their phenotypic, agronomic, historical or other cultural values for future utilization. In the present study the modified PVS2 droplet vitrification technique was tested and optimized for cryopreservation of a large diversity of blackcurrant (R. nigrum L.) accessions propagated in vitro and selected into a national gene bank core collection. Out of four accessions tested to optimize the method, three recovered and regenerated by 89–97% on average, but one recalcitrant in vitro line only by 25%. The tested post-cryopreservation recovery media with different macronutrient and growth regulator levels showed no generalized effect on regenerated shoots, but the effect of recovery media was different between cultivars. When the whole regeneration chain from cryopreservation via micropropagation to greenhouse conditions was tested, shoots at least 1 cm in length were found necessary for successful transfer ex vitro. The long-term cryopreservation of 22 blackcurrant accessions was finally conducted, with practices slightly modified from the tested protocol. The estimated recovery of shoot tips after 9 weeks in vitro was 17–94% with at least 75% recovery in seven accessions and at least 40% recovery in 19 out of 22 accessions. Only one accession had no cryopreservation success. The results demonstrated that the modified droplet vitrification technique is appropriate for a large diversity of blackcurrant accessions. However, cultivar-related differences and recovery procedures are to be considered for success in regeneration and ex vitro adaptation.

Status of introduction and conservation in barley (Hordeum vulgare L.)

Barley is important cereal crop and grown over a broad environmental range.  There is a need to collect, conserve barley genetic resources to meet the known and unknown challenges of emerging diseases and climate change. For augmentation of diverse germplasm ICAR- National Bureau of Plant Genetic Resources (NBPGR) is the nodal institute with a well-established procedure and since its establishment in 1976 and a total of 80,124 accessions in barley were introduced/imported including trait specific/ value added accessions from different countries and International Institutes.  Additionally, each year 200-3000 accessions are imported as international nurseries/ trials. International centre for Agricultural Research in Dry Areas (ICARDA) has the global mandate for improvement of barley and is doing rebreeding activities in barley. The new variability generated is made available to researchers users in form of International nurseries and trials and new form of variation in exotic material is made available through these nurseries/trials. The national Genebank  at ICAR-NBPGR conserves over eight thousand accessions of barley of which more than one thousand five hundred accessions are of exotic origin. Access to these resources is regulated by various national and international instruments related to bio-diversity and agriculture. Access to barley germplasm from India is regulated by Biological Diversity Act, 2002.  As barley is included in the multilateral system of the International Treaty on plant genetic resources for food and agriculture (ITPGRFA) and India has also designated 468 accessions to the MLS, the access to such designated accessions of barley is also made through the national focal point of the Treaty that is Department of Agricultural Cooperation and Farmers Welfare (DACFW).  However access under the provisions of the Treaty is done after signing the Standard Material Transfer Agreement (SMTA), ensuring  that the material accessed under the MLS of the treaty shall be freely available to others for use in research, breeding and training provided the third and subsequent parties are bound by the same conditions of the SMTA. Intellectual Property Rights (IPRs) cannot be claimed by the recipients on the material received from the MLS and if any commercial utilization is done, the benefits would be deposited in a trust fund of the Treaty. DOI.10.25174/2582-2675/2020/83213

Plant genetic resources in India: management and utilization.

Plant genetic resources (PGR) are the foundation of agriculture as well as food and nutritional security. The ICAR-NBPGR is the nodal institution at national level for management of PGR in India under the umbrella of Indian Council of Agricultural Research (ICAR), New Delhi. India being one of the gene-rich countries faces a unique challenge of protecting its natural heritage while evolving mutually beneficial strategies for germplasm exchange with other countries. The Bureaus activities include PGR exploration, collection, exchange, characterization, evaluation, conservation and documentation. It also has the responsibility to carry out quarantine of all imported PGR including transgenics meant for research purposes. The multifarious activities are carried out from ICAR-NBPGR headquarters and its 10 regional stations located in different agro-climatic zones of India. It has linkages with international organizations of the Consultative Group on International Agricultural Research (CGIAR) and national crop-based institutes to accomplish its mandated activities. NBPGR collects and acquires germplasm from various sources, conserves it in the Genebank, characterizes and evaluates it for different traits and provides ready material for breeders to develop varieties for farmers. ICAR-NBPGR encompasses the National Genebank Network and at present, the National Genebank conserves more than 0.40 million accessions. NBPGR works in service-mode for effective utilization of PGR in crop improvement programmes which depends mainly on its systematic characterization and evaluation, and identification of potentially useful germplasm. NBPGR is responsible for identifying trait-specific pre-adapted climate resilient genotypes, promising material with disease resistance and quality traits which the breeders use for various crop improvement programmes. The system has contributed immensely towards safeguarding the indigenous and introducing useful exotic PGR for enhancing the agricultural production. Presently, our focus is on characterization of ex situ conserved germplasm and detailed evaluation of prioritized crops for enhanced utilization; assessment of impact of on-farm conservation practices on genetic diversity; genome-wide association mapping for identification of novel genes and alleles for enhanced utilization of PGR; identification and deployment of germplasm/landraces using climate analog data; validation of trait-specific introduced germplasm for enhanced utilization. Key words: plant genetic resources; gene banks; wild relatives; biotic and abiotic stresses; marker-assisted selection.

Development of a genomic DNA reference material panel for thalassemia genetic testing.

Thalassemia is one of the most common autosomal recessive inherited diseases worldwide, and it is also highly prevalent and variable in southern China. Various types of genetic testing technologies have been developed for diagnosis and screening of thalassemia. Characterized genomic DNA reference materials (RMs) are necessary for assay development, validation, proficiency testing, and quality assurance. However, there are no publicly available RMs for thalassemia genetic testing as yet. To address the need for the publicly available DNA RMs for thalassemia genetic testing, the National Institutes for Food and Drug Control and the China National GeneBank established 32 new cell lines with three wild-type genotypes and 29 distinct genotypes of thalassemia which account for approximately 90% thalassemia carriers in China. The genomic DNA of 32 cell lines was characterized by four clinical genetic testing laboratories using different genetic testing methods and technology platforms. The genotyping results are concordant among four laboratories. In addition, the results of stability test demonstrated that the genotypes of these DNA samples are not influenced by preanalytical conditions such as long-term exposure to high-temperature (37°C) environment and repeated freeze-thawing. We developed the first national panel of 32 genomic DNA RMs which are renewable and publicly available for the quality assurance of various genetic testing methods and will facilitate research and development in thalassemia genetic testing.

Conservation of Ribes and Rubus cultivars in the Swedish National Gene Bank

Conservation of <i>Ribes</i> and <i>Rubus</i> cultivars in the Swedish National Gene Bank

Development of Laos Khao Kai Noi rice landrace (Oryza sativa L.) core collection as a model for rice genetic resources management in the Laos National Genebank

Khao Kai Noi rice is considered as an elite quality landrace in Laos, which has led to its germplasm conservation in the Laos National Genebank. As happens with other germplasm collections, a manageable yet representative sub collection has become an essential element for researchers and breeders to simplify many activities, including those related to crop improvement, phenotype-genotype correlation and determination of diversity hotspots. In this study, 109 accessions were used as a test collection for core collection development to determine the feasibility of collection reduction in a closely related rice group. Three core collections were developed by two established methodologies and evaluated by diversity indexes, allele retention, phylogenetic distribution and geographical location. Based on SSR molecular markers and PowerCore, a reduction to 24 accessions was achieved with the conservation of complete genetic diversity. A K-means based on reduction to 24 accessions rendered slightly lesser results while based on 12 accessions resulted in a 17% diversity loss. These core collections may be useful for genebank management, research and breeding activities in the future. Also, they may as well serve to estimate core collection development behavior in other landraces and cultivars, which is fundamental in genetic resources management and utilization.

Serendipitous In Situ Conservation of Faba Bean Landraces in Tunisia: A Case Study

Cultivation of faba bean (Vicia faba L.) in Tunisia is largely based on improved varieties of the crop. However, a few farmers continue to produce local cultivars or landraces. The National Gene Bank of Tunisia (NGBT) recently launched a collection project for faba bean landraces, with special focus on the regions of the North West, traditionally devoted to cultivating grain legumes, and where around 80% of the total national faba bean cultivation area is located. The seed phenotypic features of the collected samples were studied, and the genetic diversity and population structure analyzed using simple sequence repeat markers. The genetic constitution of the present samples was compared to that of faba bean samples collected by teams of the International Center for Agricultural Research in the Dry Areas (ICARDA) in the 1970s in the same region, and stored at the ICARDA gene bank. The results of the diversity analysis demonstrate that the recently collected samples and those stored at ICARDA largely overlap, thus demonstrating that over the past 50 years, little genetic change has occurred to the local faba bean populations examined. These findings suggest that farmers serendipitously applied international best practices for in situ conservation of agricultural crops.

Проявление гетерозиса у гибридов репчатого лука, полученных на стерильной основе в условиях Азербайджана

The main goal of the study was to obtain hybrids having a heterositic effect using sterile lines of onions (Allium cepa L.) applicated to conditions of Azerbaijan. We used to study a valuable source material from the collection of the National Gene Bank of Azerbaijan. As a result of the combining ability evaluation of the sterile line and the pollinator, the most promising combinations (Arzamas S-1 × Hachmaz Mest, Luganskiy S-2 × Dusti, Arzamas S-2 × Ordubad-2, Luganskiy S-1 × Kaba, Arzamas S-2 × Sabir) were revealed the greatest heterosis efficiency of yield and shelf life quality were selected. The yield of hybrids obtained with the participation of varieties of Arzamas S-1 and Arzamas S-2 and paternal varieties, compared with the Strigunovsky variety, is 17.0–78.0 % higher on average for all the years of testing. The study of the keeping capacity of F1 hybrids showed that almost all of them have a higher keeping capacity than the original varieties. The keeping capacity of hybrids especially increases, the initial varieties of which do not have long-term storage sustainability. So, bulbs of varieties Dustu, Peshpazak, Sabir, Khachmaz mestniy, Ordubad-1 for 180-200 days of storage remained only 26-29 %. Hybrids, obtained with the participation of these varieties; by the end of the study period, the keeping capacity remained at 49.8-88.7 %, i.e. 1.9-3.1 times better. Many F1 hybrids perform heterosis and early maturation, the combinations Arzamas S-2 × Strigunovsky, Arzamas S-2 × Dusti, Arzamas S-1 × Strigunovsky, Lugansky S-2 × Vostochny, Lugansky S-2 × Dusti, Arzamas S-1 × Khachmaz mestniy, Luganskiy S-1 × Kabo were more early matured than the standard for 14-18 days.

Monitoring Adventitious Presence of Transgenes in Cotton Collections from Genebank and Experimental Plots: Ensuring GM-Free Conservation and Cultivation of Genetic Resources

Bt cotton is the commercialized genetically modified (GM) crop in India with four approved GM events, namely MON531 (Bollgard® I), MON15985 (Bollgard® II), GFM and Event 1. Several GM events of cotton with insect-resistant or herbicide-tolerant GM traits were under field trials or imported for research purposes. With the increase in adoption of Bt cotton, unintentional introgression of transgenes in the conserved diversity-rich germplasm and in the field samples needs to be monitored in a systematic manner employing efficient GM diagnostics. Adventitious presence of transgenes was checked in 100 ex situ accessions of cotton being conserved in the National Genebank. These accessions belonging to Gossypium hirsutum, G. barbadense, G. arboreum, G. hirsutum × G. barbadense, represented seven Bt cotton growing states. Pentaplex PCR assay covering screening for over 90% of globally approved GM cotton events was employed, simultaneously targeting Cauliflower Mosaic Virus 35S promoter (P-35S), Agrobacterium tumefaciens nos terminator (T-nos), neomycin phosphotransferase (nptII) and phosphinothricin-N-acetyltransferase (pat) marker genes, and endogenous stearoyl acyl carrier protein desaturase (Sad1) gene as an internal control. Real-time PCR assays targeting P-35S and T-nos, with the limit of detection up to 0.01%, were employed for further confirmation. A study was also conducted on fifty cotton samples collected from the experimental plots, which were grown at distance of 50 and 10 metres from Bt cotton event MON15985. Adventitious presence of transgenes was checked using multiplex PCR assay targeting cry1Ac, cry2Ab2 and endogenous Sad1 gene to define a strategy for screening for specific GM events. Adventitious presence of transgenes was not detected in any of the tested samples based on the screening tests conducted. This strategy targeting specific GM traits would assist in GM-free cultivation of varieties in the experimental plots. The reported approach with wide GM coverage is reliable for checking adventitious presence of transgenes in genebanks, which could be applied to test conserved germplasm on a regular basis, for ensuring GM-free conservation of genetic resources.

Effect of stem gall disease on long-term germplasm preservation and quality seed production of coriander

To know the health status of crops germplasm received from various sources, seed health testing (SHT) is a routine process for long-term conservation of healthy seeds in the National Genebank at ICAR-National Bureau of Plant Genetic Resources, New Delhi. In this process, 1328 accessions of coriander were received in the Plant Quarantine Division for SHT through Germplsm Conservation Division. In the present study, SHT revealed that most of the samples of coriander in which the seeds are deformed suffer from stem gall (seed gall) disease caused by Protomycesmacrosporus. Infection of P. macrosporus greatly affects the size and quality of seeds. As a result, infected seeds become excessively larger (from 6.46x5.01 to 12.76x3.94 mm) as compared to normal seeds and the germination capacity of the seeds is also lost. Deformity in size and measurement of infected seeds compared to healthy seeds indicates susceptibility to stem gall disease. Our observation revealed that the chlamydospores present in the infected seeds survive for a long period (~ 2 years) and if such infected seeds are conserved for long term in the National Genebank, it is possible that along with the seeds, dormant chlamydospores also survive. While on the one hand such diseased seed can prove to be the carrier in spreading the pathogen from one place to another, on the other hand, market demand will be affected which will result in lower prices for the farmers cultivating coriander.

Collection, conservation and morphological characterization of Aegilops tauschii Coss. Accessions From Kashmir, India

Aegilops tauschii is an important crop wild relative of bread wheat (Triticum aestivum) and its gene pool is currently considered the most important resource for wheat improvement as potential gene donor for drought, disease and pest resistance. Aegilops tauschii subsp. tauschii grows abundantly at several places in Kashmir but is somewhat uncommon. Fourteen germplasm accessions were collected for the first time from diverse locations mainly in Budgam and Srinagar districts of Kashmir and conserved in the National Genebank. Analysis of data recorded on morphological traits of plant height, days to 50% flowering, days to 50% maturity, spike length, spikelets/spike, no. of seeds/spikelet, seed yield/plant and 100-grain weight revealed considerable variability. Significant positive association of days to flowering was observed with spike length and number of spikelets/ spike. Interestingly, spike length revealed a significant positive association with altitude of place from where accessions were originally collected. Principal component analysis revealed that first two components having Eigen values greater than one contributed nearly 60% variance. Cluster analysis grouped the accessions into three distinct groups mainly on the basis of spike length. The study provides basic information about this valuable crop wild relative and the information may help in its proper utilization.

Site identification survey in Garhwal Himalayas to establish safety duplicate of national genebank

Site identification survey in Garhwal Himalayas to establish safety duplicate of national genebank

Comprehending the Diversity in the regenerated set of Maize (Zea mays L.) Landraces Conserved in the Indian National genebank

Comprehending the Diversity in the regenerated set of Maize (Zea mays L.) Landraces Conserved in the Indian National genebank

Drawing up queries, forms and reports of the information system on plant genetic resources

Aim. To develop the National Plant Gene Pool Information System for the effective exchange of information and targeted use of this information in different research programs as well as for cooperation with regional and international organizations on the basis of intellectual property rights. Results and Discussion. This study was aimed at solving a set of interrelated objectives to compile a computer database of characteristics of the Global Collection of Agricultural Genetic Resources. The platform for this database was harmonized and systematized crop descriptors, which are the main component of the National Information System. As a part of these studies, the PGR Documentation Unit at the National Genebank of the Research Institute of Plant Genetic Resources worked at constructing data queries for presentation in the Information System forms. The tool part of the CAC-DB information system, where one can generate any query at one’s discretion by selecting data using the Query tools, served as a basis for this work. Seventeen types of queries for general use and 4 types of report templates were designed. The National Information System on Plant Genetic Resources, which is being developed at the Research Institute of Plant Genetic Resources, will make it possible to increase the efficiency of its storage, documentation, management and use through the exchange of information and germplasm, and will ensure the transition of genetic and breeding studies to a new technological level, increasing its effectiveness. Conclusions. The developed National Information System is unique and selective. It will concentrate comprehensive information on the plant gene pool of the Republic, as it is specific for Central Asia and the Southern Caucasus, having no analogues in the world.

CNSA: a data repository for archiving omics data.

With the application and development of high-throughput sequencing technology in life and health sciences, massive multi-omics data brings the problem of efficient management and utilization. Database development and biocuration are the prerequisites for the reuse of these big data. Here, relying on China National GeneBank (CNGB), we present CNGB Sequence Archive (CNSA) for archiving omics data, including raw sequencing data and its further analyzed results which are organized into six objects, namely Project, Sample, Experiment, Run, Assembly and Variation at present. Moreover, CNSA has created a correlation model of living samples, sample information and analytical data on some projects. Both living samples and analytical data are directly correlated with the sample information. From either one, information or data of the other two can be obtained, so that all data can be traced throughout the life cycle from the living sample to the sample information to the analytical data. Complying with the data standards commonly used in the life sciences, CNSA is committed to building a comprehensive and curated data repository for storing, managing and sharing of omics data. We will continue to improve the data standards and provide free access to open-data resources for worldwide scientific communities to support academic research and the bio-industry.Database URL: https://db.cngb.org/cnsa/.

Assessment of genetic integrity of Siberian wild rye (Elymus sibiricus L.) germplasm conserved ex situ and under accelerated aging using microsatellite markers

Genetic changes in Siberian wild rye (Elymus sibiricus L.) germplasm conserved ex situ and with accelerated aging were studied using microsatellite markers. Five accessions, divided and deposited in the National Medium-term Genebank of Forage Germplasm of China (NMGFC) and the National Genebank of China (NGC) since 1998, were sampled. Four recently-regenerated accessions were selected to assess genetic changes under accelerated aging conditions using a thermostat-controlled cabinet at 45 °C for 7–13 days to obtain seeds with germination rates (Gr) of approximately 80%, 50%, and < 30%. Twenty-four simple sequence repeat (SSR) markers were used to screen 70 seeds of each sample and 137 SSR alleles in a naturally-aged germplasm set and 113 SSR alleles in an accelerated aging germplasm set were scored. Loss of alleles was observed with respect to the primer pair and sample in the NMGFC compared to that in the NGC, but no significant differences were found for each accession in seeds stored in NMGFC compared to that in seeds stored in NGC. Among-sample differences were the largest contributors to total SSR variation, at 70.6% and 76.2% for naturally-aged and accelerated-aging samples, respectively. Gr did not separate samples in either germplasm set based on genetic association analysis. Prominent changes in genetic diversity were found both in naturally-aged and accelerated-aging samples, with significant decreases in Shannon’s information index and Nei’s genetic diversity for cases with Gr ≤ 30% for both germplasm sets. These results should be valuable to improve the safe conservation of forage, especially for Elymus (family Gramineae).