Ethyl Methanesulfonate Research Articles

Identification and Gene Cloning of a Brittle Culm Mutant (bc22) in Rice

The mechanical strength of rice culm, an essential factor for lodging resistance and yield maintenance, is influenced by the composition and structure of the cell wall. In this study, we characterized a rice brittle culm mutant 22 (bc22), derived from LR005 through ethyl methanesulfonate (EMS) mutagenesis. The bc22 culm exhibited increased fragility and reduced mechanical strength compared to LR005. The mutant displayed pleiotropic effects, including a shorter plant height and panicle length, a smaller grain size, and the absence of the glume hairs. Scanning electron microscopy revealed a decrease in cell density and a looser structure in the bc22 culms. Biochemical analysis demonstrated a significant increase in hemicellulose content and a marked reduction in lignin content in the culm of bc22. Genetic analysis indicated that the brittle culm trait was governed by a single recessive gene. After employing bulked segregant analysis (BSA), whole-genome resequencing, and MutMap methods, LOC_Os02g25230 was identified as the candidate gene responsible for bc22. In bc22, a point mutation from proline (Pro) to leucine (Leu) in its coding region led to the pleiotropic phenotype. A complementation test further confirmed that the missense mutation causing the proline to leucine amino acid substitution in LOC_Os02g25230 was causative of the observed bc22 phenotype. Additionally, gene expression analysis showed that BC22 had higher expression levels in the culms, leaves, and spikelets compared to the roots. Taken together, our findings indicate that BC22 is a pleiotropic gene, and the influence of BC22 on brittleness may be associated with cell wall biosynthesis in rice culm.

Augmenting tomato functional genomics with a genome-wide induced genetic variation resource.

Induced mutations accelerate crop improvement by providing novel disease resistance and yield alleles. However, the alleles with no perceptible phenotype but have an altered function remain hidden in mutagenized plants. The whole-genome sequencing (WGS) of mutagenized individuals uncovers the complete spectrum of mutations in the genome. Genome-wide induced mutation resources can improve the targeted breeding of tomatoes and facilitate functional genomics. In this study, we sequenced 132 doubly ethyl methanesulfonate (EMS)-mutagenized lines of tomato and detected approximately 41 million novel mutations and 5.5 million short InDels not present in the parental cultivar. Approximately 97% of the genome had mutations, including the genes, promoters, UTRs, and introns. More than one-third of genes in the mutagenized population had one or more deleterious mutations predicted by Sorting Intolerant From Tolerant (SIFT). Nearly one-fourth of deleterious genes mapped on tomato metabolic pathways modulate multiple pathway steps. In addition to the reported GC>AT transition bias for EMS, our population also had a substantial number of AT>GC transitions. Comparing mutation frequency among synonymous codons revealed that the most preferred codon is the least mutagenic toward EMS. The validation of a potato leaf-like mutation, reduction in carotenoids in ζ-carotene isomerase mutant fruits, and chloroplast relocation loss in phototropin1 mutant validated the mutation discovery pipeline. Our database makes a large repertoire of mutations accessible to functional genomics studies and breeding of tomatoes.

Improving French bean yield potential through induced mutagenesis using EMS and SA

IntroductionFrench bean (Phaseolus vulgaris L.) holds global significance as one of the most consumed legumes, with commercial value surpassing that of all other legume crops combined. In India, the consumption of French beans has grown steadily, especially in the North Eastern region, driven by heightened consumer interest in its nutritional benefits. Considering these factors, we initiated an induced mutagenesis program to enhance the genetic diversity of locally grown French bean genotypes, traditionally cultivated for their superior adaptability.MethodsTo achieve this, we initiated an induced mutagenesis program. Seeds from the village seed stock were subjected to treatments with varying doses of ethyl methane sulfonate (EMS) ranging from 0.1% to 0.4% and sodium azide (SA) from 0.1% to 0.4%. The objective was to increase yield potential and enhance genetic diversity.ResultsThe treatment with EMS and SA led to a non-specific, dosage-independent reduction in biophysiological characteristics in French bean mutants. Notably, the 0.4% SA treatment significantly inhibited germination and fertility, causing a decrease in chlorophyll (10.02 mg. g-1 FW) and carotenoid (1.57 mg. g-1 FW) levels. This suggests a disruption in genes associated with chlorophyll and carotenoid synthesis. However, in the M2 generation, the mutagenic treatments substantially improved yield and associated traits. The highest pod yield per plant was recorded at 79.50 gm for the 0.2% EMS treatment. A character association study revealed strong correlations (0.217 to 0.995) between pod yield and other agronomic traits.DiscussionThe results indicate that selecting mutants based on these traits in populations treated with EMS and SA can significantly increase crop yield. The 0.2% SA and 0.2% EMS M2 mutant populations exhibited the highest induced variability, making them ideal for selecting higher-yielding mutant lines for further breeding generations. The increased yields in these mutant lines, derived from a local cultivar, show promise for meeting the growing demand for French bean production through their widespread cultivation.

Transcriptomic Screening of Alternaria oxytropis Isolated from Locoweed Plants for Genes Involved in Mycotoxin Swaisonine Production.

Locoweed is a collective name for a variety of plants, such as Oxytropis and Astragalus L. When these plants are infected by some fungi or endophytes, they will produce an alkaloid (swainsonine) that is harmful to livestock. Chronic toxicity characterized by neurological disorders occurs in livestock overfed on locoweed, and swainsonine (SW) is considered a major toxic component. The mechanism of the SW synthesis of endophytic fungi from locoweed remains unknown. In order to further discover the possible synthetic pathway of SW, in this study, a mycotoxin (SW) producer, Alternaria oxytropis isolate, UA003, isolated from Locoweed plants, and its mutant were subjected to transcriptomic analyses to ascertain the genes involved in the synthesis of this toxin. Mutant strain A. oxytropis E02 was obtained by ethyl methanesulfonate (EMS) mutagenesis treatment, and the strains were sequenced with different culture times for transcriptomic analysis and screening of differentially expressed genes. The results show a highly significant (p < 0.01) increase in SW yield in the A. oxytropis E02 strain obtained by EMS mutagenesis treatment compared to A. oxytropis UA003. A total of 637 differentially expressed genes were screened by transcriptome sequencing analysis, including 11 genes potentially associated with SW biosynthesis. These genes were screened using GO and KEGG data annotation and analysis. Among the differential genes, evm.TU.Contig4.409, evm.TU.Contig19.10, and evm.TU.Contig50.48 were associated with L-lysine biosynthesis, the L-pipecolic acid pathway, and the α-aminoadipic acid synthesis pathway. This study provides new insights to elucidate the mechanism of SW synthesis of endophytic fungi in locoweed and provides data support for further exploration of A. oxytropis genomics studies.

Response surface methodology for optimization of biodiesel production by the Penicillium commune NRC 2016 and its mutants

AbstractWith increasing energy requirements and limited fuel resources, finding cheap renewable sources is essential. The objective of our current economic study is to enhance biodiesel production from the Penicillium commune, NRC 2016. Four mutagenic agents were used to improve the lipid production from P. commune NRC 2016: gamma radiation, ethyl methanesulfonate (EMS), ethidium bromide (Et Br), and sodium azide (NaN3). The Inter-Simple-Simplece Repeats molecular (ISSR) marker was used to compare the wild type of P. commune NRC 2016 and the resulting mutants. The results showed a major difference between the wild type and its mutants. Response surface methodology was used to optimize the culture conditions for lipid production by P. commune NRC 2016, and the induced mutants used hydrolysate that was produced from bagasse using Bacillus cereus 3SME. The maximum lipid content (g/l) for P. commune NRC 2016, P. commune NRC 2016–1, P. commune NRC 2016–2, and P. commune NRC 2016–3 were 2.01, 2.55, 1.71, and 2.27, respectively. Gas chromatographic analysis was used for biodiesel produced from P. commune NRC 2016 wild type, and its mutants were mainly C16–C18, which is suitable for transesterification. The physical properties, such as density, viscosity, cloud point, pour point, and cetane number, of the biodiesel from P. commune NRC 2016 and its mutants were similar to those of standard biodiesel and could be applied on a large scale. The biodiesels produced through P. commune NRC 2016 and its mutants can be utilized on a broad scale as eco-friendly products obtained from a low-cost carbon source.

Effect of Chemical and Physical Mutagens on Leaf Morphological Changes in Horse Gram (Macrotyloma Uniflorum (Lam.)

Horse gram (Macrotyloma Uniflorum (Lam.) varieties viz., Phule Sakas and Man were mutated with EMS (ethyl methane sulphonate), SA (Sodium azide) and gamma rays (G), to determine the mutagenic potency in breeding programme. Considerable frequency of leaf changes carrying plants could be observed in both the varieties of Horse gram as a result of mutagenic treatments. The frequency of leaf change carrying plants revealed an enhancing feature with the gradual rise in concentration /dose of EMS, SA and Gamma rays in both the Horse gram varieties.

Modification in the ascorbate-glutathione cycle leads to a better acclimation to high light in the rose Bengal resistant mutant of Nannochloropsis oceanica

Understanding how to adapt outdoor cultures of Nannochloropsis oceanica to high light (HL) is vital for boosting productivity. The N. oceanica RB2 mutant, obtained via ethyl methanesulfonate mutagenesis, was chosen for its tolerance to Rose Bengal (RB), a singlet oxygen (1O2) generator. Compared to the wild type (WT), the RB2 mutant showed higher resilience to excess light conditions. Analyzing the ascorbate-glutathione cycle (AGC), involving ascorbate peroxidases (APX, EC 1.11.1.11), dehydroascorbate reductase (DHAR, EC 1.8.5.1), and glutathione reductase (GR, EC 1.8.1.7), in the RB2 mutant under HL stress provided valuable insights. At 250 μmol photon m−2 s−1 (HL), the WT strain displayed superoxide anion radicals (O2▪-) and hydrogen peroxide (H2O2) accumulation, increased lipid peroxidation, and cell death compared to normal light (NL) conditions (50 μmol photon m−2 s−1). The RB2 mutant didn't accumulate O2▪- and H2O2 after HL exposure, and exhibited increased APX, DHAR, and GR activities and transcript levels compared to WT and remained consistent after HL treatment. Although the RB2 mutant had a smaller ascorbate (AsA) pool than the WT, its ability to regenerate dehydroascorbate (DHA) increased post HL exposure, indicated by a higher AsA/DHA ratio. Additionally, under HL conditions, the RB2 mutant displayed an improved glutathione (GSH) regeneration rate (GSH/GSSG ratio) without changing the GSH pool size. Remarkably, H2O2 or menadione (a O2▪- donor) treatment induced cell death in the WT strain but not in the RB2 mutant. These findings emphasize the essential role of AGC in the RB2 mutant of Nannochloropsis in handling photo-oxidative stress.

Studies on Phytochemical and Antioxidative Potential in Ocimum basilicum L. by inducing variability using Ethyl Methane Sulphonate in Uttarakhand

Studies on Phytochemical and Antioxidative Potential in Ocimum basilicum L. by inducing variability using Ethyl Methane Sulphonate in Uttarakhand

Investigations on the genotoxic potential of styrene in Fischer 344 rats using multiple endpoints.

Genotoxicity of styrene monomer was evaluated in male Fischer 344 rats using the alkaline comet assay for DNA damage, micronucleus assay for cytogenetic damage and the Pig-a assay for gene mutations. In a dose range finding (DRF) study, styrene was administered by oral gavage in corn oil for 28 consecutive days at 0, 100, 500, and 1000 mg/kg/day. The bioavailability of styrene was confirmed in the DRF by measuring its plasma levels at approximately 7- or 15-min following dosing. The 1000 mg/kg/day group exceeded the maximum tolerated dose based on body weight and organ weight changes and signs of central nervous system depression. Based on these findings, doses of 0, 100, 250, and 500 mg/kg/day (for 28 or 29 days) were selected for the genotoxicity assays. Animals were sacrificed 3-4 h after treatment on Day 28 or 29 for assessing various genotoxicity endpoints. Pig-a mutant frequencies and micronucleus frequencies were determined in peripheral blood erythrocytes. The comet assay was conducted in the glandular stomach, duodenum, liver, lung, and kidney. These studies were conducted in accordance with the relevant OECD test guidelines. Oral administration of styrene did not lead to genotoxicity in any of the investigated endpoints. The adequacy of the experimental conditions was assured by including animals treated by oral gavage with the positive control chemicals ethyl nitrosourea and ethyl methane sulfonate. Results from these studies supplement to the growing body of evidence suggesting the lack of in vivo genotoxic potential for styrene.

Genetic Screening of Factors in the Plant Protein Secretion.

The endomembrane system in plants is composed of interconnected membrane organelles that contribute to intracellular structure and function. These organelles include the endoplasmic reticulum (ER), Golgi apparatus, vacuole, trans-Golgi network, and prevacuolar compartment or multivesicular body. Through vesicle-mediated transport, secreted proteins are synthesized in the ER and subsequently transported along the secretory pathway to the vacuole or outside of cells to fulfill specialized functions. Genetic screening is a crucial method for studying plant protein secretion. It entails identifying phenotypic differences resulting from genetic mutations, such as ethyl methanesulfonate, T-DNA insertion, and RNAi, to investigate gene function and discover mutants with specific traits or gene functions. Significant progress has been achieved in the study of plant protein secretion through genetic screening. In this protocol, we provide a step-by-step guide to studying the protein secretion pathway using a genetic screen approach. We use the example of the free 1 suppressor of Arabidopsis thaliana and oil body mutants of Marchantia polymorpha. Additionally, we offer an overview of genetic screening and briefly summarize the emerging technologies in the field of protein secretion research.

Characterization of &lt;i&gt;Staphylococcus lugdunensis&lt;/i&gt; biofilms through ethyl methanesulfonate mutagenesis

&lt;p&gt;&lt;italic&gt;Staphylococcus lugdunensis&lt;/italic&gt; is a coagulase-negative species responsible for a multitude of infections. These infections often resemble those caused by the more pathogenic staphylococcal species, &lt;italic&gt;Staphylococcus aureus&lt;/italic&gt;, such as skin and soft tissue infections, prosthetic joint infections, and infective endocarditis. Despite a high mortality rate and infections that differ from other coagulase-negative species, little is known regarding &lt;italic&gt;S. lugdunensis&lt;/italic&gt; pathogenesis. The objective of this study is to identify the essential factors for biofilm formation in &lt;italic&gt;S. lugdunensis&lt;/italic&gt;. &lt;italic&gt;S. lugdunensis&lt;/italic&gt; was mutagenized through ethyl methanesulfonate (EMS) exposure, and the individual cells were separated using a cell sorter and examined for biofilm formation at 8 hr and 24 hr timepoints. Mutations that resulted in either increased or decreased biofilm formation were sequenced to identify the genes responsible for the respective phenotypes. A mutation within the &lt;italic&gt;S. lugdunensis&lt;/italic&gt; surface protein A (&lt;italic&gt;slsA&lt;/italic&gt;) gene was common among all of the low biofilm formers, thus suggesting that high expression of this protein is important in biofilm formation. However, other mutations common among the mutants with decreased biofilm formation were in the putative divalent cation transport gene, &lt;italic&gt;mgtE&lt;/italic&gt;. Conversely, a mutation in the gene that codes for the von Willebrand factor binding protein, &lt;italic&gt;vwbl&lt;/italic&gt;, was common among the mutants with increased biofilm formation. Following proteinase K treatment, a significant dispersal of the &lt;italic&gt;S&lt;/italic&gt;. &lt;italic&gt;lugdunensis&lt;/italic&gt; biofilm matrix occurred, thus confirming the presence of primarily protein-mediated biofilms; this is in agreement with previous &lt;italic&gt;S. lugdunensis&lt;/italic&gt; studies. Additionally, all low biofilm formers exhibited decreased protein levels (1.95–2.77 fold change) within the biofilm matrix, while no difference was observed with extracellular DNA (eDNA) or polysaccharides. This study presents a unique methodology to identify genes that affect biofilm formation and sheds light on &lt;italic&gt;S. lugdunensis&lt;/italic&gt; pathogenesis.&lt;/p&gt;

Chemical mutagenesis of Listeria monocytogenes for increased tolerance to benzalkonium chloride shows independent genetic underpinnings and off-target antibiotic resistance.

Listeria monocytogenes, a potentially fatal foodborne pathogen commonly found in food processing facilities, creates a significant economic burden that totals more than $2 billion annually in the United States due to outbreaks. Quaternary ammonium compounds (QACs), including benzalkonium chloride (BAC), are among the most widely used sanitizers to inhibit the growth and spread of L. monocytogenes from food processing facilities. However, resistance to QACs has been increasing in L. monocytogenes and different genetic mechanisms conferring resistance have been discovered. Here, we used ethyl methanesulfonate (EMS) to chemically mutagenize the BAC-susceptible strain, L. monocytogenes FSL-N1-304. We isolated two mutants with increased tolerance to BAC compared to the parental strain. Next, we assessed the off-target effect of increased tolerance to BAC by measuring the minimum inhibitory concentrations (MICs) of a diverse set of antibiotics, revealing that mut-1 and mut-2 displayed significantly increased resistance to fluoroquinolone antibiotics compared to the parental strain. A hemolysis assay was then used to investigate a potential correlation between BAC tolerance and virulence. Interestingly, mut-1 and mut-2 both exhibited significantly higher hemolysis percentage than the parental strain. We then sequenced the genomes of the parental strain and both mutants to identify mutations that may be involved in the increased resistance to BAC. We identified 3 and 29 mutations in mut-1 and mut-2, respectively. mut-1 contained nonsynonymous mutations in dagK (a diacylglycerol kinase), lmo2768 (a permease-encoding gene), and lmo0186 (resuscitation promoting factor). mut-2 contained a nonsense mutation in the nucleotide excision repair enzyme UvrABC system protein B encoding gene, uvrB, which likely accounts for the higher number of mutations observed. Transcriptome analysis in the presence of BAC revealed that genes related to the phosphotransferase system and internalins were up-regulated in both mutants, suggesting their significance in the BAC stress response. These two mutants provide insights into alternative mechanisms for increased BAC tolerance and could further our understanding of how L. monocytogenes persists in the food processing environment.

Molecular, morphological, and biomolecular characterization of ethyl methanesulfonate-induced mutations in Aerides odoratum, an orchid

Aerides odoratum L. is a medicinal and ornamental orchid of the Eastern Himalayas, India. Its uses in pharmacology are nought because of its rare occurrence, longer life cycle, and lack of artificial methods for large-scale propagation. Protocorms (60 days old) were treated with various concentrations of ethyl methanesulfonate ranging from 0.025% to 0.5% for different durations (2, 5, 7, and 10 days) during M0 (treatment) in vitro generation. The survival of these protocorms and the generation of shoots and roots were noted. 10–20 plantlets of each treatment showing variation in the number and weight of shoots in M0 generation (treatment generation) were maintained for three subsequent generations. M3 (third generation) axenic plantlets were hardened in the orchidarium with a 75% survival frequency. Four mutant lines (M3V1–M3V4) were established based on RAPD analysis of the leaf DNA of 300-day-old plants. M3V-1 was noted as the best among all mutant lines based on morphological analysis, whereas M3V-2 was noted as the most superior in terms of having the highest quantity of osmolytes, pigments, stomata, and phenol. M3V-3 had maximum root length, leaf breadth, and carotenoids. M3V-4 showed the viridis chlorophyll spectrum as the most distinguishing factor. Hence, this standardized protocol can be used for the genetic improvement of A. odoratum.

Sensitivity of seedling growth, fertility and yield contributing traits in rice (Oryza sativa L.) to electron beam and Ethyl Methane Sulphonate

Sensitivity of seedling growth, fertility and yield contributing traits in rice (Oryza sativa L.) to electron beam and Ethyl Methane Sulphonate

Study the influence of Mutagenic Treatment on Cell Suspension Culture of Elephantopus scaber L. for the Quantitative enhancement of Bioactive constituents and their Antioxidant potentials

In the present study, the impact of the chemical mutagen, Ethyl methanesulfonate (EMS) was evaluated on the quantitative enhancement of sesquiterpene, flavonoids (Quercetin and Rutin) and Stigmasterol compound Elephantopus scaber. The leaves of E. scaber L. were used for the callus induction. MS basal medium supplemented with different hormone combinations of 2,4-D and BAP (1:1 ratio) was used. The cell suspension culture was treated with three concentrations of EMS (0.1%, 0.2% and 0.5%) and treatment durations (1h, 2h and 3h). Mutagenesis is an important strategy to improve production of secondary metabolites using cell suspension culture. The results obtained revealed that the concentrations and treatment durations of EMS were most effective in enhancement in the content of studied metabolites. EMS treatment of 0.1% concentration was effective. The analysis was done using HPTLC technique. The highest content of Deoxyelephantopin recorded in 1h EMS treatment and 0.1% concentration compared to control sample. Quercetin, Rutin content was also high in 1h EMS treatment and 0.1% concentration compared to control sample. The Stigmasterol content was more in 2h EMS treatment and 0.1% concentration compared to control sample. The EMS treated samples showed potential scavenging activity compared to control samples IC50 value. The method was simple, precise, eco-friendly.

In-vitro intervention modifies the outcome of induced mutagenesis using gamma-ray but not ethyl methanesulphonate in cockscomb (Celosia cristata L.)

The viability of conventional vis-à-vis in-vitro mutagenesis was investigated using EMS and gamma-ray in cockscomb (Celosia cristataL.) ethyl methanesulphonate (EMS) was a better mutagen as compared to gamma-ray in terms of mutation frequency, effectiveness,efficiency, factor of effectiveness and mutation rate. The spectrum of mutants induced by EMS and the aforementioned parametersdecreased with the increase in its concentration. Moreover, the EMS treatment to dry seed was fructuous compared to the pre-soakedseeds. Although the effect of EMS treatment was similar in conventional and in-vitro mutagenesis; the gamma-ray treatment workeddifferently in these two modes of mutagenesis.

Biological Activities of Chamaecyparis lawsoniana (A.Murray bis) Parl. and Thuja plicata Donn ex D.Don Essential Oils: Toxicity, Genotoxicity, Antigenotoxicity, and Antimicrobial Activity

Essential oils (EOs) derived from evergreen conifer trees are recognized as complex sources of agents with various biological activities. In this study, the biological activities of the EOs derived from Chamaecyparis lawsoniana (A.Murray bis) Parl. (CLEO) and Thuja plicata Donn ex D.Don (TPEO) were investigated. The third instar larvae of the fruit fly Drosophila melanogaster were used as a model organism to determine the in vivo toxicity, genotoxicity, and antigenotoxicity of five different concentrations of CLEO and TPEO, ranging from 0.19% to 3%. Artemia salina was used for acute toxicity assessment and determination of LC50 after 24 h and 48 h. The antimicrobial activity of the CLEO and TPEO was tested against fourteen bacterial and two fungal strains using the microdilution method. The larvicidal activity and developmental time of D. melanogaster increased as the concentrations of the EOs rose. CLEO and TPEO produced a dose-dependent increase in DNA damage compared with the negative control. Both oils, at concentrations up to 1.5%, demonstrated the ability to reduce the genotoxic effect induced by ethyl methanesulfonate, with a reduction rate exceeding 60%. CLEO and TPEO were highly toxic against Artemia salina nauplii. The results indicate that the tested EOs act as antimicrobial agents against some pathogenic bacteria and fungi. CLEO and TPEO show promising potential as a source of antimicrobial and antigenotoxic agents and as potential biocides against pest insects and arthropods.

Inducing mutation and ascertaining lethal dosage of in vitro cultures of banana cv. Ney Poovan to ethyl methane sulfonate

In vitro mutation breeding in vegetatively propagated crops like banana offers a benefit in screening for beneficial variants in plant cells or cultured tissues. An attempt was made to induce mutants and determine the lethal dose, as it is the prerequisite to optimize the concentration and duration of the mutagen used to recover a larger population in mutation research. Shoot tip cultures were treated for 2 and 4 h at six different EMS concentrations ranging from 80 mM to 160 mM, whereas proliferating multiple shoots were exposed for 30 and 60 min at six different EMS concentrations ranging from 8 mM to 40 mM. Survival percentage, shoot length, and number of shoots reduced linearly and significantly as concentration and duration increased in both shoot tips and proliferating multiple buds. The probit curve-based analysis of mortality of treated explants revealed that the LD50 was 155.83 mM for 2 h and 113.72 mM for 4 h, respectively for shoot tip cultures, whereas for proliferating multiple buds, the LD50 value was adjusted to 39.11 mM for 30 min and 30.41 mM for 60 min. 160 mM EMS for 4 h resulted in a shorter shoot, a longer rooting duration, a lesser number of roots, and decreased root development. In proliferating multiple shoots, the smallest shoot, longest rooting duration, least number of roots, and shortest root were observed in 40 mM EMS for 60 min. Similar reductions in growth parameters were observed in proliferating multiple shoots at higher exposure to EMS for a longer duration.

A large sequenced mutant library - valuable reverse genetic resource that covers 98% of sorghum genes.

Mutant populations are crucial for functional genomics and discovering novel traits for crop breeding. Sorghum, a drought and heat-tolerant C4 species, requires a vast, large-scale, annotated, and sequenced mutant resource to enhance crop improvement through functional genomics research. Here, we report a sorghum large-scale sequenced mutant population with 9.5 million ethyl methane sulfonate (EMS)-induced mutations that covered 98% of sorghum's annotated genes using inbred line BTx623. Remarkably, a total of 610 320 mutationswithin the promoter and enhancer regions of 18 000 and 11 790 genes, respectively, can be leveraged for novel research of cis-regulatory elements. A comparison of the distribution of mutations in the large-scale mutant library and sorghum association panel (SAP) provides insights into the influence of selection. EMS-induced mutations appeared to be random across different regions of the genome without significant enrichment in different sections of a gene, including the 5' UTR, gene body, and 3'-UTR. In contrast, there were low variation density in the coding and UTR regions in the SAP. Based on the Ka /Ks value, the mutant library (~1) experienced little selection, unlike the SAP (0.40), which has been strongly selected through breeding. All mutation data are publicly searchable through SorbMutDB (https://www.depts.ttu.edu/igcast/sorbmutdb.php) and SorghumBase (https://sorghumbase.org/). This current large-scale sequence-indexed sorghum mutant population is a crucial resource that enriched the sorghum gene pool with novel diversity and a highly valuable tool for the Poaceae family, that will advance plant biology research and crop breeding.

Determination of optimal lethal dose for ethyl methane sulphonate (EMS) induced mutagenesis in castor (Ricinus communis L)

Castor, an indeterminate and perennial non-edible oilseed crop is unsuitable for mechanization over a large scale. Chemical mutagenesis using ethyl methane sulphonate (EMS) is resorted to exploit point mutations and induce monospike or synchronous maturity. In the present study, EMS mutagenesis dosage was standardized using combinations of concentration (0.5 %, 1 % and 1.5%) and duration (4, 8, 12 hours) of EMS treatment along with duration of pre-soaking with water (12 and 24 hours). Data on speed and percent germination of the seed along with root and shoot length at 7 days and 14 days after treatment indicated that 12 hours presoaking followed by 8 hours or 12 hours of treatment with 1.0% EMS treatment is the optimum lethal dose for EMS mutagenesis.