Mungbean Genome Research Articles

Characterization of TBP and TAFs in Mungbean (Vigna radiata L.) and Their Potential Involvement in Abiotic Stress Response.

The TATA-box binding protein (TBP) and TBP-associated factors (TAFs) constitute the transcription factor IID (TFIID), a crucial component of RNA polymerase II, essential for transcription initiation and regulation. Several TFIID subunits are shared with the Spt-Ada-Gcn5-acetyltransferase (SAGA) coactivator complex. Recent research has revealed the roles of TBP and TAFs in organogenesis and stress adaptation. In this study, we identified 1 TBP and 21 putative TAFs in the mungbean genome, among which VrTAF5, VrTAF6, VrTAF8, VrTAF9, VrTAF14, and VrTAF15 have paralogous genes. Their potential involvement in abiotic stress responses was also investigated here, including high salinity, water deficit, heat, and cold. The findings indicated that distinct genes exerted predominant influences in the response to different abiotic stresses through potentially unique mechanisms. Specifically, under salt stress, VrTBP, VrTAF2, and VrTAF15-1 were strongly induced, while VrTAF10, VrTAF11, and VrTAF13 acted as negative regulators. In the case of water-deficit stress, it was likely that VrTAF1, VrTAF2, VrTAF5-2, VrTAF9, and VrTAF15-1 were primarily involved. Additionally, in response to changes in ambient temperature, it was possible that genes such as VrTAF5-1, VrTAF6-1, VrTAF9-2, VrTAF10, VrTAF13, VrTAF14b-2, and VrTAF15-1 might play a dominant role. This comprehensive exploration of VrTBP and VrTAFs can offer a new perspective on understanding plant stress responses and provide valuable insights into breeding improvement.

Identification of VrNIP2-1 aquaporin with novel selective filter regulating the transport of beneficial as well as hazardous metalloids in mungbean (Vigna radiata L.)

Nodulin 26-like intrinsic protein (NIP) subfamily of aquaporins (AQPs) in plants, is known to be involved in the uptake of metalloids including boron, germanium (Ge), arsenic (As), and silicon (Si). In the present study, a thorough evaluation of 55 AQPs found in the mungbean genome, including phylogenetic distribution, sequence homology, expression profiling, and structural characterization, contributed to the identification of VrNIP2-1 as a metalloid transporter. The pore-morphology of VrNIP2-1 was studied using molecular dynamics simulation. Interestingly, VrNIP2-1 was found to harbor an aromatic/arginine (ar/R) selectivity filter formed with ASGR amino acids instead of GSGR systematically reported in metalloid transporters (NIP2s) in higher plants. Evaluation of diverse cultivars showed a high level of Si accumulation in leaves indicating functional Si transport in mungbean. In addition, heterologous expression of VrNIP2-1 in yeast revealed As(III) and GeO2 transport activity. Similarly, VrNIP2-1 expression in Xenopus oocytes confirmed its Si transport ability. The metalloid transport activity with unique structural features will be helpful to better understand the solute specificity of NIP2s in mungbean and related pulses. The information provided here will also serve as a basis to improve Si uptake while restricting hazardous metalloids like As in plants.

Molecular characterization and transcriptional analysis of VrWOX genes in mungbean [Vigna radiate (L.) Wilczek]

WUSCHEL-related homebox (WOX) gene family is a type of plant specific transcription factor, and belongs to the homeobox (HB) transcription factor superfamily. WOX genes play an important role in plant development, such as stem cell regulation and reproductive progress, and have been identified in many plant species. However, the information of mungbean VrWOX genes is limited. In this study, we identified 42 VrWOX genes in mungbean genome using Arabidopsis AtWOX genes as BLAST queries. VrWOX genes are unevenly distributed on 11 mungbean chromosomes, and chromosome 7 contains the most VrWOX genes. VrWOX genes are classified into three subgroups, the ancient group, the intermediate group and the modern/WUSCHEL group, which contains 19, 12 and 11 VrWOX members, respectively. Intraspecific synteny analysis revealed 12 VrWOX duplicated gene pairs in mungbean. Mungbean and Arabidopsis thaliana have 15 orthologous genes, and mungbean and Phaseolus vulgaris have 22 orthologous genes, respectively. The gene structure and conserved motif are different among VrWOX genes, indicating their functional diversity. The promoter regions of VrWOX genes contain different number and type of cis-acting elements, and VrWOX genes show distinct expression levels in eight mungbean tissues. Our study investigated the bioinformation and expression profiles of VrWOX genes, and provided essential information for further functional characterization of VrWOX genes.

Genome-Wide Identification, Expression Analysis, and Potential Roles under Abiotic Stress of the YUCCA Gene Family in Mungbean (Vigna radiata L.).

YUCCA, belonging to the class B flavin-dependent monooxygenases, catalyzes the rate-limiting step for endogenous auxin synthesis and is implicated in plant-growth regulation and stress response. Systematic analysis of the YUCCA gene family and its stress response benefits the dissection of regulation mechanisms and breeding applications. In this study, 12 YUCCA genes were identified from the mungbean (Vigna radiata L.) genome and were named based on their similarity to AtYUCCAs. Phylogenetic analysis revealed that the 12 VrYUCCAs could be divided into 4 subfamilies. The evidence from enzymatic assays in vitro and transgenetic Arabidopsis in vivo indicated that all the isolated VrYUCCAs had biological activity in response to IAA synthesis. Expression pattern analysis showed that functional redundancy and divergence existed in the VrYUCCA gene family. Four VrYUCCAs were expressed in most tissues, and five VrYUCCAs were specifically highly expressed in the floral organs. The response toward five stresses, namely, auxin (indole-3-acetic acid, IAA), salinity, drought, high temperatures, and cold, was also investigated here. Five VrYUCCAs responded to IAA in the root, while only VrYUCCA8a was induced in the leaf. VrYUCCA2a, VrYUCCA6a, VrYUCCA8a, VrYUCCA8b, and VrYUCCA10 seemed to dominate under abiotic stresses, due to their sensitivity to the other four treatments. However, the response modes of the VrYUCCAs varied, indicating that they may regulate different stresses in distinct ways to finely adjust IAA content. The comprehensive analysis of the VrYUCCAs in this study lays a solid foundation for further investigation of VrYUCCA genes' mechanisms and applications in breeding.

Endogenous hydrogen sulfide homeostasis is responsible for the difference in osmotic stress tolerance in two cultivars of Vigna radiate

Hydrogen sulfide (H 2 S) has been proved to possess many biological functions and it is an essential component of plant osmotic signaling. However, how endogenous H 2 S homeostasis is maintained and how this function determines mung bean osmotic tolerance is less explored. To fully answer this question, two cultivars with different osmotic stress tolerance (namely MM015 and MM047) were used in this study. Under osmotic stress, osmosis-tolerant cultivar MM047 had higher H 2 S-producing capacity, which was impaired in osmosis-sensitive cultivar MM015. In MM015, alleviation of the osmotic stress-induced wilting phenotype and destruction of redox homeostasis were achieved by the application of NaHS (a H 2 S donor) at both low (10 μM) and high (200 μM) concentrations. Moreover, NaHS treatment mitigated osmotic stress-induced reduction on photosynthetic capacity in MM015, as evaluated by net photosynthetic rate and Rubisco carboxylation activity, etc. However, in MM047, a similar protective role against osmotic stress-triggered damage was observed only when NaHS was applied at low concentration, whereas aggravated at high dose. Importantly, we identified three genes encoding L-cysteine desulfhydrase (VrLCDs, EC 4.4.1.1) from mung bean genome, all of which possess H 2 S-producing enzymatic activities. The molecular function of VrLCD2, a major basal and inducible VrLCD form was subsequently investigated through A.rhizogenes -induced hairy roots system. Overexpression of this gene mitigated osmotic stress-triggered oxidative damage in osmosis-sensitive cultivar, which was exacerbated osmosis-sensitive cultivar. Taken together, our results demonstrated VrLCD-modulated endogenous H 2 S homeostasis regulates mung bean osmotic tolerance through maintaining redox homeostasis and photosynthetic capacity. These results further provide valuable resource for molecular breeding and genetic development of stress-resilient mung bean crop through the modulation of endogenous H 2 S homeostasis. • Mung bean osmotic stress tolerance was associated with the regulation of endogenous H 2 S homeostasis. • The maintenance of H 2 S homeostasis mitigated the osmotic stress-induced destruction of redox homeostasis and photosynthetic capacity in mung bean. • The mung bean genome contains at least three genes encoding L-cysteine desulfhydrase • Osmotic stress tolerance was enhanced by overexpression of VrLCD2 in mung bean osmosis-sensitive cultivar, whereas exacerbated in osmosis-tolerant cultivar.

Genetic control of the lateral petal shape and identity of asymmetric flowers in mungbean (Vigna radiata L.).

Broad diversity of flowers in Fabaceae provides a good system to investigate development and evolution of floral symmetry in higher plants. Many studies have demonstrated a conserved mechanism controlling development of zygomorphic flower during last decades. However, the molecular basis of how asymmetric flower established is largely unknown. In this study, we characterized mutants named keeled wings (kw) in mungbean (Vigna radiata L.), which is a legume species with asymmetric flowers. Compared to those in the wild type plants, the lateral petals were ventralized in the kw mutants. Map-based cloning showed that KW was VrCYC3 gene in mungbean, the ortholog of Lotus japonicus CYC3 (LjCYC3) and Pisum sativum CYC3 (PsCYC3). In addition, another two CYC-like genes named VrCYC1 and VrCYC2 were identified from mungbean genome. The three CYC-like genes displayed distinct expression patterns in dorsal, lateral and ventral petals. It was found that VrCYC3 was located in nucleus. Further analysis showed that VrCYC3 had transcription activity and could interact with VrCYC1 and VrCYC2 in yeast cell. Moreover, the deletion of two amino acid residues in the R domain of VrCYC3 protein could decrease its interaction with VrCYC1 and VrCYC2 proteins. Our results suggest that LjCYC3/VrCYC3 orthologs play conserved roles determining the lateral petal shape and identity of zygomorphic flower as well as asymmetric flower in Papilionoideae.

Thirty Years of Mungbean Genome Research: Where Do We Stand and What Have We Learned?

Mungbean is a socioeconomically important legume crop in Asia that is currently in high demand by consumers and industries both as dried beans and in plant-based protein foods. Marker-assisted and genomics-assisted breeding are promising approaches to efficiently and rapidly develop new cultivars with improved yield, quality, and resistance to biotic and abiotic stresses. Although mungbean was at the forefront of research at the dawn of the plant genomics era 30 years ago, the crop is a “slow runner” in genome research due to limited genomic resources, especially DNA markers. Significant progress in mungbean genome research was achieved only within the last 10 years, notably after the release of the VC1973A draft reference genome constructed using next-generation sequencing technology, which enabled fast and efficient DNA marker development, gene mapping, and identification of candidate genes for complex traits. Resistance to biotic stresses has dominated mungbean genome research to date; however, research is on the rise. In this study, we provide an overview of the past progress and current status of mungbean genomics research. We also discuss and evaluate some research results to provide a better understanding of mungbean genomics.

Construction of High-Density Genetic Map and Identification of a Bruchid Resistance Locus in Mung Bean (Vigna radiata L.).

Mung bean (Vigna radiata L.) is an economically important grain legume cultivated in Asian countries. High-density genetic linkage is a valuable and effective tool for mapping quantitative trait loci (QTL). In the current study, a high-resolution genetic map containing 4,180 single-nucleotide polymorphisms (SNPs) was assigned to 11 linkage groups (LGs) and spanning 1,751.39 cM in length was constructed for mung bean, and the average distance between adjacent markers was 0.42 cM. Bruchids (Callosobruchus spp.) cause significant damage to and loss of legume seeds. A locus for bruchid resistance was detected. The gene Vradi05g03810, encoding a probable resistance-specific protein, was found to be the most likely key candidate gene in mung beans. A 69-bp sequence deletion was identified in the coding region by comparing the cDNA sequences of bruchid-resistant and bruchid-susceptible lines. This SNP-based high-density linkage map is one of the first to be constructed across the mung bean genome. This map will not only facilitate the genetic mapping of genes or complex loci that control important agronomic traits but also offer a tool for promoting future genetics and comparative genomic studies in Vigna.

Genome-wide identification and characterization of mungbean CIRCADIAN CLOCK ASSOCIATED 1 like genes reveals an important role of VrCCA1L26 in flowering time regulation

BackgroundCIRCADIAN CLOCK ASSOCIATED 1 like (CCA1L) proteins are important components that participate in plant growth and development, and now have been characterized in multiple plant species. However, information on mungbean CCA1L genes is limited.ResultsIn this study, we identified 27 VrCCA1L genes from the mungbean genome. VrCCA1L genes were unevenly distributed on 10 of the 11 chromosomes and showed one tandem and two interchromosomal duplication events. Two distinct kinds of conserved MYB domains, MYB 1 and MYB 2, were found, and the conserved SHAQK(Y/F) F sequence was found at the C terminus of each MYB 2 domain. The VrCCA1Ls displayed a variety of exon-intron organizations, and 24 distinct motifs were found among these genes. Based on phylogenetic analysis, VrCCA1L proteins were classified into five groups; group I contained the most members, with 11 VrCCA1Ls. VrCCA1L promoters contained different types and numbers of cis-acting elements, and VrCCA1Ls showed different expression levels in different tissues. The VrCCA1Ls also displayed distinct expression patterns under different photoperiod conditions throughout the day in leaves. VrCCA1L26 shared greatest homology to Arabidopsis CCA1 and LATE ELONGATED HYPOCOTYL (LHY). It delayed the flowering time in Arabidopsis by affecting the expression levels of CONSTANS (CO), FLOWERING LOCUS T (FT), and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1).ConclusionWe identified and characterized 27 VrCCA1L genes from mungbean genome, and investigated their spatio-temporal expression patterns. Further analysis revealed that VrCCA1L26 delayed flowering time in transgenic Arabidopsis plants. Our results provide useful information for further functional characterization of the VrCCA1L genes.

Genome-Wide Analysis of Late Embryogenesis Abundant Protein Gene Family in Vigna Species and Expression of VrLEA Encoding Genes in Vigna glabrescens Reveal Its Role in Heat Tolerance.

Late embryogenesis abundant (LEA) proteins are identified in many crops for their response and role in adaptation to various abiotic stresses, such as drought, salinity, and temperature. The LEA genes have been studied systematically in several crops but not in Vigna crops. In this study, we reported the first comprehensive analysis of the LEA gene family in three legume species, namely, mung bean (Vigna radiata), adzuki bean (Vigna angularis), and cowpea (Vigna unguiculata), and the cross-species expression of VrLEA genes in a wild tetraploid species, Vigna glabrescens. A total of 201 LEA genes from three Vigna crops were identified harboring the LEA conserved motif. Among these 55, 64, and 82 LEA genes were identified in mung bean, adzuki bean, and cowpea genomes, respectively. These LEA genes were grouped into eight different classes. Our analysis revealed that the cowpea genome comprised all eight classes of LEA genes, whereas the LEA-6 class was absent in the mung bean genome. Similarly, LEA-5 and LEA-6 were absent in the adzuki bean genome. The analysis of LEA genes provides an insight into their structural and functional diversity in the Vigna genome. The genes, such as VrLEA-2, VrLEA-40, VrLEA-47, and VrLEA-55, were significantly upregulated in the heat-tolerant genotype under stress conditions indicating the basis of heat tolerance. The successful amplification and expression of VrLEA genes in V. glabrescens indicated the utility of the developed markers in mung bean improvement. The results of this study increase our understanding of LEA genes and provide robust candidate genes for future functional investigations and a basis for improving heat stress tolerance in Vigna crops.

Mungbean (Vigna radiata (L.) R. Wilczek): Progress in Breeding and Future Challenges

Mungbean (Vigna radiata (L.) R. Wilczek) is a short duration farmer preferred warm-season pulse crop. The crop has shown a balanced growth worldwide, especially in developing countries. Mungbean being a great source of protein with higher folate and iron levels attracts high demand and price on the market making the farmers happy and satisfied. Moreover, it can fix atmospheric nitrogen through symbiosis with nitrogen-fixing bacteria, making it perfect for rice based cropping systems and intercropping with other crops. Despite having so many benefits, mungbean has been a neglected crop compared to other pulses with limited efforts aiming at its breeding and development. Higher productivity, breeding for biotic and abiotic stresses resistance and nutritional quality improvement are future challenges for mungbean breeders. Several researchers are working in the direction of collecting and maintaining mungbean genetic resources. Mutation breeding and genetic engineering has also been enhanced in mungbean varietal improvement. Genomic information is limited compared to other legume species. However, the recent successful sequencing of mungbean genome has opened new vistas into the crop’s R&D. It is a self-pollinated pulse with small genome size, which could be used as a model for studying other legumes. Mungbean breeders at present times aim to identify useful alleles from diverse germplasm and markers closely associated with desirable traits. The high-throughput marker genotyping system has now made it feasible to pinpoint the exact gene locations and mutations contributing target phenotypes. In this review we present the current status of conventional and molecular breeding of the crop and summary of efforts in the utilization of genetic information and genomic resources for further mungbean improvement.

Analysis of genetic variability in F2 interspecific hybrids of mung bean (Vigna radiata) using inter-retrotransposon amplified polymorphism marker system

Abstract. Fatmawati Y, Setiawan AB, Purwantoro A, Respatie DW, Teo CH. 2021. Analysis of genetic variability in F2 interspecific hybrids of mung bean (Vigna radiata) using inter-retrotransposon amplified polymorphism marker system. Biodiversitas 22: 4880-4889. Mung bean (Vigna radiata L. Wilczek) categorized as one of the pivotal annual crops of Vigna genera is commonly cultivated in rotation with the cereal crops during the drought season. Conversely, to ameliorate its stunted productivity, the interspecific hybridization technique has been introduced between the mung bean and the common bean to promote genetic improvement with the breeding projects in Indonesia. However, since mung bean is a self-pollinated crop and has a narrow genetic base, the selection and improvement of a specific trait using marker-assisted selection is more challenging. Hence, a precautionary investigation is imperative to evaluate the progenies resulting from interspecific hybridization using an ideal marker. This study aimed to investigate the genetic variability of the F2 population of the interspecific mung bean hybrids using retrotransposon-based markers, particularly Inter-Retrotransposon Amplified Polymorphism (IRAP) markers. In this study, we identified retrotransposon from the mung bean genome and determined the Long Terminal Repeat (LTR) sequence using the LTR Finder. The IRAP primers were designed from a conserved region of the LTR sequence. One hundred of the F2 interspecific hybrids generated from the crossing between mung bean and common bean were successfully discriminated by IRAP markers. The IRAP marker showed high heterozygosity and moderate Polymorphic Information Content (PIC) values. The IRAP markers were able to detect genetic variability in the F2 progenies resulting from the interspecific hybridization. Cluster analysis showed that 100 of the F2 progenies were grouped into three clusters. This study demonstrated that retrotransposon-based markers can offer an effective approach for evaluating the segregation in the F2 population of intercross hybrids in the mung bean.

Genome-wide analysis of OSCA gene family members in Vigna radiata and their involvement in the osmotic response

BackgroundMung bean (Vigna radiata) is a warm-season legume crop and belongs to the papilionoid subfamily of the Fabaceae family. China is the leading producer of mung bean in the world. Mung bean has significant economic and health benefits and is a promising species with broad adaptation ability and high tolerance to environmental stresses. OSCA (hyperosmolality-gated calcium-permeable channel) gene family members play an important role in the modulation of hypertonic stress, such as drought and salinity. However, genome-wide analysis of the OSCA gene family has not been conducted in mung bean.ResultsWe identified a total of 13 OSCA genes in the mung bean genome and named them according to their homology with AtOSCAs. All the OSCAs were phylogenetically split into four clades. Phylogenetic relationship and synteny analyses showed that the VrOSCAs in mung bean and soybean shared a relatively conserved evolutionary history. In addition, three duplicated VrOSCA gene pairs were identified, and the duplicated VrOSCAs gene pairs mainly underwent purifying selection pressure during evolution. Protein domain, motif and transmembrane analyses indicated that most of the VrOSCAs shared similar structures with their homologs. The expression pattern showed that except for VrOSCA2.1, the other 12 VrOSCAs were upregulated under treatment with ABA, PEG and NaCl, among which VrOSCA1.4 showed the largest increased expression levels. The duplicated genes VrOSCA2.1/VrOSCA2.2 showed divergent expression, which might have resulted in functionalization during subsequent evolution. The expression profiles under ABA, PEG and NaCl stress revealed a functional divergence of VrOSCA genes, which agreed with the analysis of cis-acting regulatory elements in the promoter regions of VrOSCA genes.ConclusionsCollectively, the study provided a systematic analysis of the VrOSCA gene family in mung bean. Our results establish an important foundation for functional and evolutionary analysis of VrOSCAs and identify genes for further investigation of their ability to confer abiotic stress tolerance in mung bean.

Development of Genome-wide Simple Sequence Repeat Markers from Whole-genome Sequence of Mungbean (Vigna radiata)

Background: Mungbean is an important pulse crop and it is mainly cultivated in Asia for human consumption. Its small genome and diploid nature make it a well-suited model organism among legume crops. Thus, cost-effective, reliable and highly polymorphic molecular markers distributing the whole genome are needed for diversity, mapping and functional genomics studies in this model species. Methods: The whole-genome sequence of mungbean was obtained and used as a source of identification of simple sequence repeats (SSR). The sequence reads were aligned and SSRs detection was performed using the Phobos plugin tandem repeat finder in the Geneious software program. A total of 12 mungbean genotypes were selected to validate the newly developed SSR markers. Result: In the present study, a total of 12, 49,774 and 11, 86, 386 perfect and imperfect SSR repeats were identified from the mungbean genome. The tri-repeats were the most abundant (26.10%), followed by hexa (20.82%), penta (20.45%), tetra (17.65%) and di-repeats (14.95%). We designed 1330 SSR primers based on the genomic sequence of flanking perfect SSRs (Di and tri-repeats). Among them, 50 SSR primers uniformly distributed across the 11 mungbean chromosomes were selected and used to validate 12 mungbean genotypes. The newly developed genomic SSR markers generated in the present study are a valuable genomic resource for the mungbean breeding programs.

Identification and characterization of SNPs in released, landrace and wild accessions of mungbean (Vigna radiata (L.) Wilczek) using whole genome re-sequencing

Mungbean [Vigna radiata (L.) R. Wilczek var. radiata] is vital grain legume having nutritional and socio-economic importance, especially in the developing countries. We performed whole-genome re-sequencing of three accessions representing the wild progenitor species, released and landrace of mungbean to identify SNPs with relevance to genetic relationships analysis. Approximately 9.3 million raw reads were obtained using Ion Torrent PGM™ platform and more than 92% of the reads were mapped to the reference mungbean genome. We identified a total of 233,799 single-nucleotide polymorphisms in relation to the reference genome (SNPs: 103,341 in wild, 93,078 in released and 37,380 in landrace accessions) and 9544 insertions and deletions (InDels: 4742 in wild, 3608 in released and 1194 in landrace accessions) in the coding and non-coding regions. In all accessions, genomic variants were unevenly distributed within and across the mungbean chromosomes. Among these 5339, 4739 and 1795 SNPs were non-synonymous in 815, 790 and 317 genes of wild, released and landrace accessions, respectively. These polymorphisms might contribute to the variation in important pathways of genes for abiotic and biotic stress tolerance and important agronomic traits, such as seed dormancy, flowering time and seed size in mungbean. Among the randomly selected SNPs, a selected subset was validated using Sanger sequencing technique. The genomic variations among mungbean wild, released and landrace accessions constitute a powerful tool to support genetic research and molecular breeding of mungbean.

Candidate gene mapping reveals VrMLO12 (MLO Clade II) is associated with powdery mildew resistance in mungbean (Vigna radiata [L.] Wilczek)

Loss/reduction of function of Mildew Locus O (MLO) genes clade V and MLO clade IV has been shown to be responsible for powdery mildew (PM) resistance in several plant species. Mungbean (Vigna radiata) genome possesses 18 MLO genes, VrMLO1 - VrMLO18. A previous study using mungbean F2 and BC1F1 populations derived from a cross between “CN60″ (susceptible) and “RUM5″ (resistance) demonstrated that QTL qPMRUM5−3 is a major QTL for PM resistance caused by Erysiphe polygoni and is the same with major QTL qPMV4718−3 that confers PM resistance in “V4718″ (resistance). In this study, bioinformatics analysis revealed VrMLO12 locates in the qPMRUM5−3 region. Fine mapping in the F2 and BC1F1 populations using newly developed DNA markers including gene-specific markers demonstrated association between VrMLO12 and the PM resistance. Sequence analyses of VrMLO12 revealed that compared to susceptible mungbeans, RUM5 and V4718 possess SNPs in exon 10 and exon 13. The SNPs caused amino acid changes of VrMLO12, A387S and A476 G, respectively. The change occurred in transmembrane 6 domain and calmodulin binding domain (CaMBD) of the VrMLO12 protein, respectively. qRT-PCR showed that transcript expression level of VrMLO12 in RUM5 challenged with and without by E. polygoni was significantly higher than that in CN60. Phylogenetic analysis revealed that in contrast to previous findings that MLO proteins associated with PM resistance belong to MLO clade V and MLO clade IV, VrMLO12 belongs to MLO clade II. The result suggested that VrMLO12 may function differently from the other MLOs that associated with PM susceptibility. Our findings provide insight into the PM resistance in mungbean and tools for mungbean breeding.

Submergence response of pyruvate decarboxylase family genes in adzuki bean

Pyruvate decarboxylase (PDC) is the main enzyme in ethanol fermentation in plant low-oxygen response. Expression of the PDC gene can be induced by submergence, a low-oxygen state, which has been demonstrated in Arabidopsis, maize and rice among other plants. However, it is unknown whether the gene expression has similar pattern in the Fabaceae family under low-oxygen stress, and unavailability of PDC family member sequences in Fabaceae hindered these studies. We aimed to comprehensively identify PDC genes in adzuki bean, and characterize their phylogenetic relationships and expression profile with the emphasis of expression response to submergence. Based on genome sequences and identified conserved domains, HMMER was used to mine PDC genes in adzuki bean in addition to mung bean, both of which are Vigna species under the Fabaceae family. Sequences were aligned using MUSCLE and Phylogenetic topology was decided by using MrBayes. Tissue expression patterns were decided using publically available RNASeq data. Adzuki bean plants were treated with submergence stress, and expressions under treatments were measured using qRT-PCR. We identified the genes in PDC family in the genomes of adzuki bean and mung bean. The known PDC peptides contain three conserved domains with the same order. Based on the domain composition, we identified four PDC family genes in adzuki bean, and three in mung bean. Phylogenetic reconstruction resolved three major classes. Class I of the family was grouped with PDC gene in Arabidopsis, and thus likely encodes PDC; one gene is present in this class in adzuki bean, whose expression was high in roots and nodules compared to other tissues, and was significantly induced under submergence treatment, reaching highest at 12 h after waterlogging. No Class I gene was identified in mung bean, possibly result of assembly gaps or failure of gene model prediction of mung bean genome. Class II genes are clustered together with larger subunit of acetolactate synthase, which have two representatives in mung bean and adzuki bean, respectively. There is one copy of gene in Class III in the genome of the two Vigna species, respectively, which is grouped with an uncharacterized Arabidopsis gene. Interestingly, the adzuki bean gene in this class was significantly induced by waterlogging, whose expression reaching the highest earlier than the Class I gene, which indicates that both the Class I and Class III genes in the PDC family are involved in low-oxygen stress response in adzuki bean. Pyruvate decarboxylase family genes were mined in adzuki bean and mung bean, whose phylogeny was clarified into three independent classes. Expression of a putative PDC gene was significantly elevated by submergence stress in adzuki bean. The Class III gene in adzuki bean was significantly induced in expression by waterlogging, too, yet its expression reached highest earlier than putative PDC genes, suggesting involvement of the Class III genes in low-oxygen stress response.

Genome-wide identification and expression analysis of SnRK2 gene family in mungbean (Vigna radiata) in response to drought stress

Drought stress causes lower crop production globally. Plants have acquired many adaptations to overcome drought stress. Mungbean (Vigna radiata (L.) R.Wilczek) is a legume crop widely cultivated in South, East and Southeast Asia. It is grown in high-temperature areas where drought is the main cause of reduced plant growth and productivity. Plants cope with drought stress by activating different signalling mechanisms. The sucrose non-fermenting-1-related protein kinase 2 family (SnRK2s) is known to play vital roles in osmotic stress and in abscisic acid (ABA) signalling pathways by phosphorylating downstream targets. The genes encoding SnRK2s in mungbean and their detailed characterisation remain unexplored. We have conducted extensive genome-wide analysis for gene prediction, in silico gene analysis, evolutionary analysis and gene-expression profiling under drought-stress conditions by quantitative real-time PCR. Through genome-wide analysis, eight SnRK2 genes were predicted in the mungbean genome and were assigned the names VrSnRK2.1–VrSnRK2.8, according to their order on the chromosomes. The VrSnRK2 genes identified were classified into three clusters based on their phylogenetic relationship with those of Arabidopsis thaliana. Drought stress was imposed on 11-day-old mungbean plants by completely withholding water for 3 days. According to real-time qPCR data, the expression of most of the VrSnRK2 genes was induced by drought stress, indicating their role in the drought-stress response. One of the genes, namely SnRK2.6c, showed highest expression level (12-fold) under drought stress, possibly indicating a critical role under water-deficit conditions. These data provide important information about the VrSnRK2 gene family in mungbean. The results will help in future functional characterisation of VrSnRK2 genes.

Effect of Processing on Mungbean (Vigna radiata) Flour Nutritional Properties and Protein Composition

Pulses are traditionally processed prior to consumption, providing opportunities for modifying nutritional composition, dependant on the type of pulse and method used. In this study, we investigated the effect of whole seed, dehulling (dahl), germination and roasting on changes in mungbean flour nutritional properties, protein composition and relative protein abundance. Processed flours were analysed and compared for protein content, moisture, fat, ash, dietary fibre, total starch and amylose. Significant differences were imparted on dietary fibre content, with roasting and germination increasing the ratio of insoluble/soluble fibre as well as resistant starch. Comparative proteomic analysis resulted in a combined total of 539 protein identifications, searching against the Mungbean reference genome (NCBI Vigna radiata Annotation Release 100). Normalised spectral abundance factors were used as a measure of relative abundance and statistical analysis was applied (Students’ T-Test), where proteins with a p-value of < 0.05 considered significantly different. Processing imparted considerable changes to nutritional composition and should be further exploited for food applications. The comparative proteomic analyses carried out in this study proved useful for investigating the effect of processing on subsequent changes in protein composition and relative abundance.

Genome-wide identification of AP2/ERF transcription factors in mungbean (Vigna radiata) and expression profiling of the VrDREB subfamily under drought stress

Mungbean (Vigna radiata L.) is a valuable legume crop grown in tropical and subtropical areas of Asia. Drought is one of the major factors hindering its growth globally. APETALA2/ethylene-responsive element factor binding proteins (AP2/ERF) are an important family of plant-specific transcription factors (TFs) involved in drought-stress tolerance. We identified 71 AP2/ERF TFs in the mungbean genome by using bioinformatics tools and classified them into subfamilies: AP2 (16 members), ERF (22), RAV (2), DREB (30) and soloist (other proteins with no domain, 1). Members of DREB play a critical role in drought-stress tolerance. Ten-day-old mungbean plants cv. AZRI-06 were exposed to drought stress by complete withholding of water for 7 days. Root samples were collected from control and drought-stressed plants, and the expression pattern of 30 identified VrDREB genes was determined by qPCR. Most VrDREB genes exhibited differential expression in response to drought. Five genes (VrDREB5, VrDREB12, VrDREB13, VrDREB22, VrDREB30) were highly expressed under drought stress and might be considered excellent candidates for further functional analysis and for improvement of mungbean drought tolerance.