Pea Gene Research Articles

Bio-engineered As-Ag-TiO2 nanoparticles enhance the genetic transformation of Pisum sativum L. via proton-coupled electron transfer-dependent alternative protonation

Nanoparticle-assisted biomolecule delivery to improve plant gene transformation and expression is a, cutting-edge approach. In this study, we examined the effects of acetosyringone-doped BENPs on the enhanced genetic transformation of peas. The vir gene inducer acetosyringone was delivered into Agrobacterium via a nano-doping system with TiO2 and Ag NPs. Four BENPs, such as TiO2 (S1), Ag-TiO2 (S2), As-TiO2 (S3), and As-Ag-TiO2 (S4), were green-synthesized from the PLE using hydrothermal and calcination approaches. SEM, HR-TEM-EDAX, SAED, XRD, and XPS physicochemical characterizations ensured the BENPs structure and biomolecule doping. The potential bar transformants were chosen against the MIC of BASTA® using the EHA 105 Agrobacterium tumefaciens strain carrying the pCAMBIA 1304-plasmid. The delivery of nano-doped acetosyringone was studied with various applications, and the typical exogenous acetosyringone delivery in the absence of BENPs was also analyzed as a control experiment. The gus histochemical, PCR, and southern hybridization confirmed that, among the BENPs, S4 at a 4 ppm dose induced the highest gene transformation efficiency (36.33 %) in pea along with three days of co-cultivation, 30 sec of sonication, and 3 min of vacuum infiltration. In BENP-assisted pea gene transformation, S2 at 4 ppm (26.66 %), S3 at 3 ppm (23 %), and S1 at 4 ppm (17.66 %) showed the next three significant gene transformation efficiencies, respectively. The elevated virC and virG gene expression profiles of the S4 BENP-treated Agrobacterium were identified with gene-specific RT-PCR analysis. The PCET activity of S4 was critically confirmed with cyclic voltammetry, which revealed that BENPs might function as an alternative proton supplement system to improve the vir gene induction and transformation of T-DNA. Hence, this alternative PCET-dependent BENP-assisted acetosyringone delivery for Agrobacterium mediated gene transformation exhibits potential value in the development of genetically-modified peas and opens the door to crops for advanced genetic manipulation against various biotic and abiotic stresses.

Genome-wide identification, structural characterization and gene expression analysis of the WRKY transcription factor family in pea (Pisum sativum L.)

BackgroundThe WRKY gene family is one of the largest families of transcription factors in higher plants, and WRKY transcription factors play important roles in plant growth and development as well as in response to abiotic stresses; however, the WRKY gene family in pea has not been systematically reported.ResultsIn this study, 89 pea WRKY genes were identified and named according to the random distribution of PsWRKY genes on seven chromosomes. The gene family was found to have nine pairs of tandem duplicates and 19 pairs of segment duplicates. Phylogenetic analyses of the PsWRKY and 60 Arabidopsis WRKY proteins were performed to determine their homology, and the PsWRKYs were classified into seven subfamilies. Analysis of the physicochemical properties, motif composition, and gene structure of pea WRKYs revealed significant differences in the physicochemical properties within the PsWRKY family; however, their gene structure and protein-conserved motifs were highly conserved among the subfamilies. To further investigate the evolutionary relationships of the PsWRKY family, we constructed comparative syntenic maps of pea with representative monocotyledonous and dicotyledonous plants and found that it was most recently homologous to the dicotyledonous WRKY gene families. Cis-acting element analysis of PsWRKY genes revealed that this gene family can respond to hormones, such as abscisic acid (ABA), indole-3-acetic acid (IAA), gibberellin (GA), methyl jasmonate (MeJA), and salicylic acid (SA). Further analysis of the expression of 14 PsWRKY genes from different subfamilies in different tissues and fruit developmental stages, as well as under five different hormone treatments, revealed differences in their expression patterns in the different tissues and fruit developmental stages, as well as under hormone treatments, suggesting that PsWRKY genes may have different physiological functions and respond to hormones.ConclusionsIn this study, we systematically identified WRKY genes in pea for the first time and further investigated their physicochemical properties, evolution, and expression patterns, providing a theoretical basis for future studies on the functional characterization of pea WRKY genes during plant growth and development.

Identification and expression analysis of SBP-Box-like (SPL) gene family disclose their contribution to abiotic stress and flower budding in pigeon pea (Cajanus cajan).

The SPL gene family (for Squamosa Promoter-binding like Proteins) represents specific transcription factors that have significant roles in abiotic stress tolerance, development and the growth processes of different plants, including initiation of the leaf, branching and development of shoot and fruits. The SPL gene family has been studied in different plant species; however, its role is not yet fully explored in pigeon pea (Cajanus cajan ). In the present study, 11 members of the CcSPL gene family were identified in C. cajan . The identified SPLs were classified into nine groups based on a phylogenetic analysis involving SPL protein sequences from C. cajan , Arabidopsis thaliana , Cicer arietinum , Glycine max , Phaseolus vulgaris , Vigna unguiculata and Arachis hypogaea . Further, the identification of gene structure, motif analysis, domain analysis and presence of cis -regulatory elements in the SPL family members were studied. Based on RNA-sequencing data, gene expression analysis was performed, revealing that CcSPL2.1, 3 and 13A were significantly upregulated for salt-tolerance and CcSPL14 and 15 were upregulated in a salt-susceptible cultivar. Real-time qPCR validation indicated that CcSPL3, 4, 6 and 13A were upregulated under salt stress conditions. Therefore, molecular docking was performed against the proteins of two highly expressed genes (CcSPL3 and CcSPL14 ) with three ligands: abscisic acid, gibberellic acid and indole-3-acetic acid. Afterward, their binding affinity was obtained and three-dimensional structures were predicted. In the future, our study may open avenues for harnessing CcSPL genes in pigeon pea for enhanced abiotic stress resistance and developmental traits.

Real-time expression and in silico characterization of pea genes involved in salt and water-deficit stress

Real-time expression and in silico characterization of pea genes involved in salt and water-deficit stress

Genome-wide identification, expression analysis, and functional study of the bZIP transcription factor family and its response to hormone treatments in pea (Pisum sativum L.)

BackgroundBasic leucine zipper (bZIP) protein is a plant-specific transcription factor involved in various biological processes, including light signaling, seed maturation, flower development, cell elongation, seed accumulation protein, and abiotic and biological stress responses. However, little is known about the pea bZIP family.ResultsIn this study, we identified 87 bZIP genes in pea, named PsbZIP1 ~ PsbZIP87, via homology analysis using Arabidopsis. The genes were divided into 12 subfamilies and distributed unevenly in 7 pea chromosomes. PsbZIPs in the same subfamily contained similar intron/exon organization and motif composition. 1 tandem repeat event and 12 segmental duplication events regulated the expansion of the PsbZIP gene family. To better understand the evolution of the PsbZIP gene family, we conducted collinearity analysis using Arabidopsis thaliana, Oryza sativa Japonica, Fagopyrum tataricum, Solanum lycopersicum, Vitis vinifera, and Brachypodium distachyon as the related species of pea. In addition, interactions between PsbZIP proteins and promoters containing hormone- and stress-responsive cis-acting elements suggest that the regulation of PsbZIP expression was complex. We also evaluated the expression patterns of bZIP genes in different tissues and at different fruit development stages, all while subjecting them to five hormonal treatments.ConclusionThese results provide a deeper understanding of PsbZIP gene family evolution and resources for the molecular breeding of pea. The findings suggested that PsbZIP genes, specifically PSbZIP49, play key roles in the development of peas and their response to various hormones.

Understanding the structure and function of GDSL-type esterase/lipase genes in pigeon pea for their role under moisture stress conditions

The SGNH hydrolase superfamily's GDSL-type esterase/lipase proteins (GELPs) play a crucial role in plant growth and stress tolerance. However, limited knowledge exists regarding the molecular role of GELP genes under drought conditions, especially in pigeon pea. In this study, 112 GDSL genes were found, distributed over the 11 pigeon pea chromosomes, and classified into 5 clades based on the phylogenetic analysis. The pigeon pea GDSL gene family seems to have grown significantly as a result of tandem duplications (48 %). Five exons and four introns represent the coding areas of more than 71.4 % of the CcGDSL genes, according to our analysis. According to the cis-elements study, CcGDSL gene expression was observed to be mostly influenced by fluctuating stressful situations and also by light and plant hormones. We could narrow down to 14 stress-responsive CcGDSL genes in order to assess their expression levels under seedling-stage drought stress conditions. Our data indicates that among these 112 genes, CcGDSL21, CcGDSL63, CcGDSL64 and CcGDSL92 could be involved in regulating the primary response under drought stress. This preliminary study will add to our understanding of GDSL genes in pigeon pea and help in elucidating their role in drought tolerance in pigeon pea.

Identification, characterization, and validation of NBS-encoding genes in grass pea.

Grass pea is a promising crop with the potential to provide food and fodder, but its genomics has not been adequately explored. Identifying genes for desirable traits, such as drought tolerance and disease resistance, is critical for improving the plant. Grass pea currently lacks known R-genes, including the nucleotide-binding site-leucine-rich repeat (NBS-LRR) gene family, which plays a key role in protecting the plant from biotic and abiotic stresses. In our study, we used the recently published grass pea genome and available transcriptomic data to identify 274 NBS-LRR genes. The evolutionary relationships between the classified genes on the reported plants and LsNBS revealed that 124 genes have TNL domains, while 150 genes have CNL domains. All genes contained exons, ranging from 1 to 7. Ten conserved motifs with lengths ranging from 16 to 30 amino acids were identified. We found TIR-domain-containing genes in 132 LsNBSs, with 63 TIR-1 and 69 TIR-2, and RX-CCLike in 84 LsNBSs. We also identified several popular motifs, including P-loop, Uup, kinase-GTPase, ABC, ChvD, CDC6, Rnase_H, Smc, CDC48, and SpoVK. According to the gene enrichment analysis, the identified genes undergo several biological processes such as plant defense, innate immunity, hydrolase activity, and DNA binding. In the upstream regions, 103 transcription factors were identified that govern the transcription of nearby genes affecting the plant excretion of salicylic acid, methyl jasmonate, ethylene, and abscisic acid. According to RNA-Seq expression analysis, 85% of the encoded genes have high expression levels. Nine LsNBS genes were selected for qPCR under salt stress conditions. The majority of the genes showed upregulation at 50 and 200μM NaCl. However, LsNBS-D18, LsNBS-D204, and LsNBS-D180 showed reduced or drastic downregulation compared to their respective expression levels, providing further insights into the potential functions of LsNBSs under salt stress conditions. They provide valuable insights into the potential functions of LsNBSs under salt stress conditions. Our findings also shed light on the evolution and classification of NBS-LRR genes in legumes, highlighting the potential of grass pea. Further research could focus on the functional analysis of these genes, and their potential use in breeding programs to improve the salinity, drought, and disease resistance of this important crop.

Genome-wide identification, characterization, and validation of the bHLH transcription factors in grass pea.

Background: The basic helix-loop-helix (bHLH) transcription factor is a vital component in plant biology, with a significant impact on various aspects of plant growth, cell development, and physiological processes. Grass pea is a vital agricultural crop that plays a crucial role in food security. However, the lack of genomic information presents a major challenge to its improvement and development. This highlights the urgency for deeper investigation into the function of bHLH genes in grass pea to improve our understanding of this important crop. Results: The identification of bHLH genes in grass pea was performed on a genome-wide scale using genomic and transcriptomic screening. A total of 122 genes were identified as having conserved bHLH domains and were functionally and fully annotated. The LsbHLH proteins could be classified into 18 subfamilies. There were variations in intron-exon distribution, with some genes lacking introns. The cis-element and gene enrichment analyses showed that the LsbHLHs were involved in various plant functions, including response to phytohormones, flower and fruit development, and anthocyanin synthesis. A total of 28 LsbHLHs were found to have cis-elements associated with light response and endosperm expression biosynthesis. Ten conserved motifs were identified across the LsbHLH proteins. The protein-protein interaction analysis showed that all LsbHLH proteins interacted with each other, and nine of them displayed high levels of interaction. RNA-seq analysis of four Sequence Read Archive (SRA) experiments showed high expression levels of LsbHLHs across a range of environmental conditions. Seven highly expressed genes were selected for qPCR validation, and their expression patterns in response to salt stress showed that LsbHLHD4, LsbHLHD5, LsbHLHR6, LsbHLHD8, LsbHLHR14, LsbHLHR68, and LsbHLHR86 were all expressed in response to salt stress. Conclusion: The study provides an overview of the bHLH family in the grass pea genome and sheds light on the molecular mechanisms underlying the growth and evolution of this crop. The report covers the diversity in gene structure, expression patterns, and potential roles in regulating plant growth and response to environmental stress factors in grass pea. The identified candidate LsbHLHs could be utilized as a tool to enhance the resilience and adaptation of grass pea to environmental stress.

Aquaporin genes in garden pea and their regulation by the nano-antioxidant fullerol in imbibing embryos under osmotic stress

Aquaporins (AQPs) are known as small membrane intrinsic proteins that help to transport water and certain solutes through biological membranes. AQPs gene families are being extensively studied in major crops, but less investigated in pea (<italic>Pisum sativum</italic> L.), which is an economically significant legume crop with a huge complex genome. Here, we present a genome-wide identification, structural characterization, subcellular localization, and expression profiling of the AQPs in pea with a particular interest in their involvement in nano fullerol-conferred osmotic stress alleviation. We identified 39 full-length aquaporin genes from the pea genome, which were classified into five subfamilies. The protein structure of aquaporins appears to have substrate-specific residues which are conserved in plants, allowing for inference of substrate specificity. In particular, the PsNIP2-2-2 was identified with a Gly-Ser-Gly-Arg (GSGR) selective filter that indicates the ability to uptake silicon. Analysis of tissue transcriptomes revealed preferred expressions of certain <italic>PsAQPs</italic> in the underground, aerial and reproductive organs, respectively. Development-regulated expressions of two <italic>PsTIPs</italic> and two <italic>PsPIPs</italic> in seeds were noticed. RNA-Seq of the imbibing embryos treated with mannitol (M) or mannitol plus 100 mg/L fullerol (MF) revealed two <italic>PsTIPs</italic> being similarly regulated by M or MF, three <italic>PsNIPs</italic> being up-regulated only by M without F, and four other genes that were only regulated under MF condition. To our knowledge, this is the first report on transcriptional regulation of AQPs by fullerols, which adds to our knowledge on the plant-carbon nano-substances interactions.

Pea transformation: History, current status and challenges

This review recapitulates the history, important milestones, the current status, and the perspectives of the pea (Pisum sativum L.) transformation as a tool for pea crop breeding. It summarises the developments of the pea transformation from the first methodological experiments to achieving the complete transformation and regeneration of genetically modified (GM) plants, transformation with the first genes of interest (GOI), to recent techniques of targeted genome editing. We show how recent biotechnological methods and genetic engineering may contribute to pea breeding in order to speed up the breeding process and for the creation of new pea cultivars. The focus is laid on genetic engineering which represents an excellent technology to enhance the pea gene pool with genes of interest which are not naturally present in the pea genome. Different methods of pea transformation are mentioned, as well as various GOI that have been used for pea transformation to date, all aimed at improving transgenic pea traits. Tolerance to herbicides or resistance to viruses, fungal pathogens, and insect pests belong, among others, to the pea traits that have already been modulated by methods of genetic engineering. The production of phytopharmaceuticals is also an important chapter in the use of genetically modified peas. We compare different methods of introducing transgenes to peas and also the usage of different selective and reporter genes. The transformation of other major legumes (soybeans, beans) is marginally mentioned. The effect of genetically modified (GM) peas on animal health (feeding tests, allergenicity) is summarised, the potential risks and benefits of pea modification are evaluated and also the prime expectations of GM peas and the real current state of this technology are compared. Unfortunately, this technology or, more precisely, the products created by this technology are under strict (albeit not scientifically-based) legislative control in the European Union.

Genome-wide transcriptome analysis and characterization of the cytochrome P450 flavonoid biosynthesis genes in pigeon pea (Cajanus cajan).

226 CcCYP450 genes were identified at the genomic level and were classified into 45 clades based on phylogenetic analysis. CcCYP75B165 gene was found that might play important roles in the biosynthesis of flavonoids in pigeon pea, and was significantly induced by methyl jasmonate (MeJA). The cytochrome P450 mono-oxygenase (CYP450) superfamily plays a key role in the flavonoid biosynthesis pathway and resists different kinds of stresses. Several CYP450 genes have been identified to be involved in the biosynthesis of crop protection agents. However, the CcCYP450 genes from pigeon pea have not been identified. Here, 226 CcCYP450 genes were identified at the genomic level by analysing the gene structure, distribution on chromosomes, gene duplication, and conserved motifs and were classified into 45 clades based on phylogenetic analysis. RNA-seq analysis revealed clear details of CcCYP450 genes that varied with time of MeJA (methyl jasmonate) induction. Among them, six CcCYP450 subfamily genes were found that might play important roles in the biosynthesis of flavonoids in pigeon pea. The overexpression of CcCYP75B165 in pigeon pea significantly induced the accumulation of genistin and downregulated the contents of cajaninstilbene acid, apigenin, isovitexin, and genistein and the expression of flavonoid synthase genes. This study provides theoretical guidance and plant genetic resources for cultivating new pigeon pea varieties with high flavonoid contents and exploring the molecular mechanisms of the biosynthesis of flavonoids under MeJA treatment.

Defining the components of the miRNA156-SPL-miR172 aging pathway in pea and their expression relative to changes in leaf morphology

The timing of developmental phase transitions is crucial for plant reproductive success, and two microRNAs (miRNA), miR156 and miR172, are implicated in the control of these changes, together with their respective SQUAMOSA promoter binding–like (SPL) and APETALA2-like (AP2) targets. While their patterns of regulation have been studied in a growing range of species, to date they have not been examined in pea (Pisum sativum), an important legume crop and model species. We analysed the recently released pea genome and defined nine miR156, 21 SPL, four miR172, and five AP2-like genes. Phylogenetic analysis of the SPL genes in pea, Medicago and Arabidopsis confirmed the eight previously defined clades, and identified a ninth potentially legume-specific SPL clade in pea and Medicago. Among the PsSPL, 14 contain a miR156 binding site and all five AP2-like transcription factors in pea include a miR172 binding site. Phylogenetic relationships, expression levels and temporal expression changes identified PsSPL2a/3a/3c/6b/9a/9b/13b/21, PsmiR156d/j and PsmiR172a/d as the most likely of these genes to participate in phase change in pea. Comparisons with leaf morphology suggests that vegetative phase change is unlikely to be definitively marked by a change in leaflet number. In addition, the timing of FT gene induction suggests that the shift from the juvenile to the adult vegetative phase may occur within fourteen days in plants grown under inductive conditions, and calls into question the contribution of miR172/AP2 to the floral transition. This work provides the first insight into the nature of vegetative phase change in pea, and an important foundation for future functional studies.

Laser Microdissection of Pisum sativum L. Nodules Followed by RNA-Seq Analysis Revealed Crucial Transcriptomic Changes during Infected Cell Differentiation

Garden pea (Pisum sativum L.) is a globally important legume crop. Like other legumes, it forms beneficial symbiotic interactions with the soil bacteria rhizobia, gaining the ability to fix atmospheric nitrogen. In pea nodules, the meristem is long-lasting and results in the formation of several histological zones that implicate a notable differentiation of infected host cells. However, the fine transcriptional changes that accompany differentiation are still unknown. In this study, using laser microdissection followed by RNA-seq analysis, we performed transcriptomic profiling in the early infection zone, late infection zone, and nitrogen fixation zone of 11-day-old nodules of pea wild-type line SGE. As a result, a list of functional groups of differentially expressed genes (DEGs) in different nodule histological zones and a list of genes with the most prominent expression changes during nodule development were obtained. Their analyses demonstrated that the highest amount of DEGs was associated with the nitrogen fixation zone. Among well-known genes controlling nodule development, we revealed genes that can be novel players throughout nodule formation. The characterized genes in pea were compared with those previously described in other legumes and their possible functions in nodule development are discussed.

Genome-wide analysis and characterization of R2R3-MYB family in pigeon pea (Cajanus cajan) and their functional identification in phenylpropanoids biosynthesis.

Thirty CcMYB were identified to involve in flavonoid and lignin biosynthesis in pigeon pea genome. A comprehensive analysis of gene structure, phylogenetic relationships, distribution on chromosomes, gene duplication, and expression patterns was performed. MYB transcription factor is one of the largest gene families in plants and plays critical roles in plant growth and development, as well as resistance to biotic and abiotic stress. However, the function of MYB genes in pigeon pea (Cajanus cajan) remains largely unknown. Here, 30 R2R3-MYB which involved flavonoid and lignin biosynthesis were identified in the pigeon pea genome and were classified into five groups based on phylogenetic analysis. Simultaneously, another 122 key enzyme genes from biosynthetic pathways of flavonoid and lignin were identified and all of them were mapped on 11 chromosomes with the co-linearity relationship. Among these genes, the intron/exon organization and motif compositions were conserved and they have undergone a strong purifying selection and tandem duplications during evolution. Expression profile analysis demonstrated most of these genes were expressed in different tissues and responded significantly to MeJA, RNA-seq analysis revealed clear details of genes varied with time of induction. Ten key genes from the phenylpropanoid pathway were selected to further verify whether they responded to induction under different abiotic stress conditions (UV-B, cold, heat, salt, drought, and GA3). This study elaborates on potential regulatory relationships between R2R3-MYB genes and some key genes involved in flavonoid and lignin biosynthesis under MeJA treatment, as well as adding to the understanding of improving abiotic stress tolerance and regulating the secondary metabolism in woody crops. A simplified discussion model for the different regulation networks involved with flavonoid and lignin biosynthesis in pigeon pea is proposed.

Dynamical Modeling of the Core Gene Network Controlling Transition to Flowering in Pisum sativum.

Transition to flowering is an important stage of plant development. Many regulatory modules that control floral transition are conservative across plants. This process is best studied for the model plant Arabidopsis thaliana. The homologues of Arabidopsis genes responsible for the flowering initiation in legumes have been identified, and available data on their expression provide a good basis for gene network modeling. In this study, we developed several dynamical models of a gene network controlling transition to flowering in pea (Pisum sativum) using two different approaches. We used differential equations for modeling a previously proposed gene regulation scheme of floral initiation in pea and tested possible alternative hypothesis about some regulations. As the second approach, we applied neural networks to infer interactions between genes in the network directly from gene expression data. All models were verified on previously published experimental data on the dynamic expression of the main genes in the wild type and in three mutant genotypes. Based on modeling results, we made conclusions about the functionality of the previously proposed interactions in the gene network and about the influence of different growing conditions on the network architecture. It was shown that regulation of the PIM, FTa1, and FTc genes in pea does not correspond to the previously proposed hypotheses. The modeling suggests that short- and long-day growing conditions are characterized by different gene network architectures. Overall, the results obtained can be used to plan new experiments and create more accurate models to study the flowering initiation in pea and, in a broader context, in legumes.

Identification of BELL Transcription Factors Involved in Nodule Initiation and Development in the Legumes Pisum sativum and Medicago truncatula.

Single three-amino acid loop extension (TALE) homeodomain proteins, including the KNOTTED-like (KNOX) and BEL-like (BELL) families in plants, usually work as heterodimeric transcription factor complexes to regulate different developmental processes, often via effects on phytohormonal pathways. Nitrogen-fixing nodule formation in legumes is regulated by different families of homeodomain transcription factors. Whereas the role of KNOX transcription factors in the control of symbiosis was studied early, BELL transcription factors have received less attention. Here, we report the identification and expression analysis of BELL genes in the legume plants Medicago truncatula and Pisum sativum, which are involved in regulating symbiosis initiation and development. A more precise analysis was performed for the most significantly upregulated PsBELL1-2 gene in pea. We found that the PsBELL1-2 transcription factor could be a potential partner of PsKNOX9. In addition, we showed that PsBELL1-2 can interact with the PsDELLA1 (LA) protein-regulator of the gibberellin pathway, which has a previously demonstrated important role in symbiosis development.

Symbiotic Regulatory Genes Controlling Nodule Development in Pisum sativum L.

Analyses of natural variation and the use of mutagenesis and molecular-biological approaches have revealed 50 symbiotic regulatory genes in pea (Pisum sativum L.). Studies of genomic synteny using model legumes, such as Medicago truncatula Gaertn. and Lotus japonicus (Regel) K. Larsen, have identified the sequences of 15 symbiotic regulatory genes in pea. These genes encode receptor kinases, an ion channel, a calcium/calmodulin-dependent protein kinase, transcription factors, a metal transporter, and an enzyme. This review summarizes and describes mutant alleles, their phenotypic manifestations, and the functions of all identified symbiotic regulatory genes in pea. Some examples of gene interactions are also given. In the review, all mutant alleles in genes with identified sequences are designated and still-unidentified symbiotic regulatory genes of great interest are considered. The identification of these genes will help elucidate additional components involved in infection thread growth, nodule primordium development, bacteroid differentiation and maintenance, and the autoregulation of nodulation. The significance of symbiotic mutants of pea as extremely fruitful genetic models for studying nodule development and for comparative cell biology studies of legume nodules is clearly demonstrated. Finally, it is noted that many more sequences of symbiotic regulatory genes remain to be identified. Transcriptomics approaches and genome-wide sequencing could help address this challenge.

Screening of CBL genes in pigeon pea with focus on the functional analysis of CBL4 in abiotic stress tolerance and flavonoid biosynthesis

Pigeon pea, the only woody legume in the world,is also a medicinal-food homogenous crop. At the same time, the calcineurin B-like (CBL) proteins that mediate a variety of abiotic stress responses are largely unknown. Here, we performed a bioinformatics analysis of the CBL family in pigeon pea, and initially verified the function of CcCBL4 in stress resistance and secondary metabolic synthesis. CBL family genes respond to 4 °C, 42 °C, NaCl, and drought stresses, and we observed the chlorophyll, relative water, proline and soluble sugar in CcCBL4-OE (over-expression) were significantly higher than CK (empty vectors) plants after stresses, and the integrity of the cell membrane was better. Seven secondary metabolites were detected, and we found that the p-coumaric acid in OE was 1.4-fold that in CK plants, but the apigenin and pterostilbene were reduced. And the expression of genes in these pathways was also confirmed changes. Altogether, our work revealed a potential regulation mechanism between CcCBL proteins, stress resistance and flavonoid metabolism, which is conducive to enhance the adaptability of pigeon pea to the environment.

Diversity Analysis and Identification of Promising Powdery Mildew Resistance Genotypes in Field Pea (&amp;lt;i&amp;gt;Pisum sativum&amp;lt;/i&amp;gt; L.)

In the present study, seventy-one field pea gene pools including three released varieties were evaluated in an augmented block design for assessing genetic divergence and level of resistance to powdery mildew for exploitation in a breeding program aimed at improving yield potential of field pea by using cluster and principal component analysis. Among the 10 studied traits, four (Eigenvalue >1.0) contributed more than 68.45% variability among the materials. Cluster analysis grouped the 71 field pea genotypes into seven distinct classes. The genetic divergence between all possible pairs of clusters were highly significant (P<0.01). The inter-cluster D² value ranged from 311.63 to 2850.61 indicated that the evaluated gene pools were highly divergent. The genetically more divergent materials present in cluster five and six as indicated by inter-cluster distance value (2850.61). Selecting genotypes of these clusters and crossing them probably provide promising recombinants and better sergeants for future breeding program. Considerable variation was also found for resistance against the powdery mildew diseases. Out of the total 71 genotypes 12 were resistant, 29 were moderately resistant, 25 were moderately susceptible and 5 were susceptible to powdery mildew disease. Among 12 resistant genotypes; GPHA-9 and GPHA-19 were high yielder and GPHA-29, GPHA-48, GPHA-45 and GPHA-42 genotypes were found to be high yielding among 29 moderately resistant genotypes. The resistant genotypes identified could be exploited directly and/or may be transferred through hybridization to high yielding disease susceptible genotypes after checking their yield and disease stability in a number of locations and seasons for more confirmation with the present finding, since the present result was from one location and one season (year) data.

New sources of Sym2A allele in the pea (Pisum sativum L.) carry the unique variant of candidate LysM-RLK gene LykX.

At the onset of legume-rhizobial symbiosis, the mutual recognition of partners occurs based on a complicated interaction between signal molecules and receptors. Bacterial signal molecules named Nod factors (“nodulation factors”) are perceived by the plant LysM-containing receptor-like kinases (LysM-RLKs) that recognize details of its structure (i.e., unique substitutions), thus providing the conditions particular to symbiosis. In the garden pea (Pisum sativum L.), the allelic state of Sym2 gene has long been reported to regulate the symbiotic specificity: for infection to be successful, plants with the Sym2A allele (for “Sym2 Afghan”, as these genotypes originate mostly from Afghanistan) require an additional acetylation of the Nod factor which is irrelevant for genotypes with the Sym2E allele (for “Sym2 European”). Despite being described about 90 years ago, Sym2 has not yet been cloned, though phenotypic analysis suggests it probably encodes a receptor for the Nod factor. Recently, we described a novel pea gene LykX (PsLykX) from the LysM-RLK gene family that demonstrates a perfect correlation between its allelic state and the symbiotic specificity of the Sym2A-type. Here we report on a series of Middle-Eastern pea genotypes exhibiting the phenotype of narrow symbiotic specificity discovered in the VIR plant genetic resources gene bank (Saint-Petersburg, Russia). These genotypes are new sources of Sym2A, as has been confirmed by an allelism test with Sym2A pea cv. Afghanistan. Within these genotypes, LykX is present either in the allelic state characteristic for cv. Afghanistan, or in another, minor allelic state found in two genotypes from Tajikistan and Turkmenistan. Plants carrying the second allele demonstrate the same block of rhizobial infection as cv. Afghanistan when inoculated with an incompatible strain. Intriguingly, this “Tajik” allele of LykX differs from the “European” one by a single nucleotide polymorphism leading to an R75P change in the receptor part of the putative protein. Thus, our new data are in agreement with the hypothesis concerning the identity of LykX and the elusive Sym2 gene.