Showing papers on "Mutation breeding published in 2021"

A highly mutable GST is essential for bract colouration in Euphorbia pulcherrima Willd. Ex Klotsch

Abstract: Mutation breeding is an extraordinary tool in plant breeding to increase the genetic variability, where mutations in anthocyanin biosynthesis are targets to generate distinctive phenotypes in ornamental species. In poinsettia, ionizing radiation is routinely applied in breeding programs to obtaining a range of colours, with nearly all pink and white varieties being obtained after γ- or X-ray mutagenesis of red varieties. In the present study we performed a thorough characterization of a potential mutagenesis target gene as the main responsible for the ‘white paradox’ in poinsettia. We identified a GST gene in poinsettia (Bract1) as an essential factor for the expression of anthocyanin-based red colouration of bracts, which presents a high phylogenetic similarity to known anthocyanin-related GSTs. Red poinsettia varieties and white mutants generated from these varieties by X-ray were analysed for polymorphisms related to the ‘white paradox’ in the species. A 4 bp mutation in a short repeat within the coding region of Bract1 is most likely responsible for the appearance of white phenotypes upon irradiation treatment. The polymorphism between wild-type and mutant alleles co-segregates with the phenotype in progeny from heterozygous red and white parents. Moreover, overexpression of Bract1 wild-type allele in Arabidopsis tt19 mutants restored the anthocyanin phenotype, while the Bract1 mutated allele showed to be non-functional. The identified repeat seems to be highly unstable, since mutated plants can be easily detected among fewer than 200 shoots derived from 10 mutated plants. Our data indicate that particular short repeat sequences, similar to microsatellite sequences or so-called dynamic mutations, might be hot spots for genetic variability. Moreover, the identification of the Bract1 mutation fills a gap on the understanding on the molecular mechanism of colour formation in poinsettia.

Development of Novel Blackgram (Vigna mungo (L.) Hepper) Mutants and Deciphering Genotype × Environment Interaction for Yield-Related Traits of Mutants

Abstract: Blackgram (Vigna mungo (L.) Hepper) yields are noticeably poor due to a shortage of improved varieties and an aggravated narrow genetic base. An attempt was made to isolate novel blackgram mutants by selecting for yield-related traits derived through gamma irradiation and testing the mutant genotype’s stability across the different environments. The irradiated blackgram populations M1-M5 were established in the background of cultivars ADT 3, Co 6, and TU 17-9. Desirable mutants were selected from M3 to M5 generations. It was observed in M2 and M3 that gamma rays showed higher mutagenic efficacy and generated good inherited variance for the yield-related traits. M4 established three divergent groups in each blackgram cultivar revealed by clustering analysis. The number of pods per plant, number of clusters per plant, and number of pods per cluster showed a strong direct association with single plant yield and could be considered as selection traits. G × E interactions were higher than the variation due to genotype for single plant yield. Limited environmental interaction was observed for the genotypes G24, G16, G36, G30, and G17, as revealed by AMMI, and the genotypes G18 and G29, as revealed by GGE. GGE biplot revealed the environment-specific genotypes G13 for E1 (Aduthurai), G7 for E2 (Kattuthottam), and G34 for E3 (Vamban) and also portrayed the highly discriminating (E3) and representative (E2) environments. Selected novel blackgram genotypes from this research are useful genetic stocks for genetic improvement and breeding.

Induced mutagenesis for genetic improvement of Allium genetic resources: a comprehensive review

Abstract: The genus Allium is one of the largest monocotyledon genera having 1000 designated species with commercial and economic significance. Allium cepa L. (onion) and Allium sativum L. (garlic) are the most imperative edible species of this genus which are cultivated and consumed globally. Despite that, not much systematic and focused research has been carried out in these crops due to constraints like biennial nature, high crossability, cross or sexual incompatibilities/limited sexual reproduction, obligate apomict, large genome size, high heterozygosity, etc. To create genetic variability, induced mutagenesis is the optimum and best alternative for Alliums especially for the development of genetically improved and wider adaptive cultivars under changing climatic scenario. Development of new cultivars having tolerance against various biotic and abiotic stresses needs continuous efforts and attention of the breeder. For that, sufficient and wide genetic variation in the germplasm is the driving force for the breeders to select best genotypes. Development of mutants can be an alternative breeding strategy since mutations cause heritable genetic variations, which provide the eventual foundation for the evolution of new cultivars, forms or species. Such variations could be created artificially through various chemicals or physical agents, known as mutagens. Mutation breeding is an efficient and conventional method of crop improvement. With the aid of modern omics and molecular markers, allium breeding could be accelerated to develop desired products under rapidly climate scenario. This is the first comprehensive and detailed review on induced mutagenesis and mutation breeding in alliums.

Effects of Carbon Ion Beam Irradiation on Phenotypic Variations and Biochemical Parameters in Early Generations of Soybean Plants

Abstract: Understanding the characteristics of carbon ion beam irradiation-induced mutation is essential to its potential application in plant breeding. A carbon ion beam-mutagenized soybean population was generated from the newly released soybean variety Dongsheng 28, with irradiation dosages of 100 Gy, 120 Gy and 140 Gy. Many phenotypic variations and novel mutants with heritable tendencies including plant height mutants, sterile mutants, early mature mutants, rolled leaves and short petioles mutants, yield-related mutants and lodging-resistant mutants were identified. Diverse variations in seed size, seed protein and oil concentration were found. Increasing irradiation dosage from 100 Gy to 140 Gy increased leaf chlorophyll concentration in M1 generation, but this effect was significantly reduced in M2 generation. The activities of superoxide dismutase (SOD), peroxidase (POD) and malondialdehyde (MDA) concentration all showed wider variation in M1 and M2 generation, the only exception being that the MDA concentration was similar to the control in the M2 generation. Overall, we suggest that treating soybean seeds with carbon ion beam irradiation at a dosage of 120 Gy (80 Mev/u) could be effective in soybean mutation breeding.

Genotype-by-Sequencing Analysis of Mutations and Recombination in Pepper Progeny of Gamma-Irradiated Gametophytes.

Abstract: The irradiation of dry seeds is the most widely-used irradiation method for improving seed-propagated crops; however, the irradiation of other tissues also has useful effects. The irradiation of plant reproductive organs, rather than seeds, for mutation breeding has advantages, such as producing non-chimeric progeny. However, the mutation frequency and spectrum produced using this method have not been analyzed on a genome-wide level. We performed a genotype-by-sequencing analysis to determine the frequencies of single-base substitutions and small (1-2 bp) insertions and deletions in hot pepper (Capsicum annuum L.) plants derived from crosses using gamma-irradiated female or male gametophytes. The progeny of irradiated gametophytes showed similar or higher DNA mutation frequencies, which were dependent on the irradiation dose and irradiated tissue, and less biased single base substitutions than progeny of irradiated seeds. These characteristics were expected to be beneficial for development of mutation population with a high frequency of small DNA mutations and performing reverse-genetics-based mutation screening. We also examined the possible use of this irradiation method in manipulating the meiotic recombination frequency; however, no statistically significant increase was detected. Our results provide useful information for further research and breeding using irradiated gametophytes.

Chronic Gamma Irradiation Changes Phenotype and Gene Expression Partially Transmitted to Next-Generation Tomato Seedlings

Abstract: Compared to the studies on acute irradiation of seeds, fewer studies have reported on the chronic irradiation of seedlings, especially in fruit-bearing vegetables. We examined the effects of chronic gamma irradiation on tomato (Solanum lycopersicum ‘Micro-Tom’) seedlings exposed to gamma rays (50, 100, 150, and 200 Gy) for 4 weeks. As the total dose of gamma rays increased, leaf length, trichome density, and seed number were reduced in the irradiated seedlings (M1). Additionally, a change in fruit shape was observed. Chronic gamma irradiation reduced the expression of two trichome-related genes and affected the expression levels of 11 reactive oxygen species (ROS)-related genes. We examined the transmittance of these effects using M2 plants. The trichome density and fruit shape were similar between M2 and control plants; however, a reduction in leaf length and seed number was detected in M2 plants. Interestingly, changes in the expression of four ROS-related genes (ZAT10, Mn-SOD, POD3, and RBOH1) found in M1 were detected in M2 plants. Thus, the changes in phenotype and gene expression induced by chronic gamma irradiation were transmitted to the next generation. Additionally, we found novel mutants from M2 plants, suggesting that chronic gamma irradiation may be considered in tomato mutation breeding.

Development of High Yielding Cowpea [Vigna unguiculata (L.) Walp.] Lines with Improved Quality Seeds through Mutation and Pedigree Selection Methods

Abstract: Cowpea [Vigna unguiculata (L.) Walp.] is a major legume crop and an important source of protein in Africa. The Kafr El-Sheikh University has a long history of cowpea breeding and improvement in Egypt. Two superior lines with high seed yield and quality were selected through mutation breeding and released to farmers as new varieties under the names Kafr El Sheikh-1 and Kaha-1. Crosses were made between these two varieties to further improve cowpea to meet farmers’ demand. Using the pedigree selection method, 13 new superior F10 lines were selected and evaluated over 2 years for seed yield and related traits, earliness, and protein content under low (16 plants/m2) and high (24 plants/m2) plant densities. The results showed that plants grown in narrower space produced significantly higher seed yield per unit area than the plants grown in wider space. All developed lines produced significantly higher seed yield than the two parental lines in the 2018 trial and Kaha-1 in the 2019 trial. Line number 6 proved to be the best genotype for earliness (73.5–73.9 days after sowing), seed yield (573–647 g/m2), and crude protein content (22.7–24.3%) in both trials. In addition, line 4 with bushy determinate growth habit and high seed quality was recently released as a new variety (Sakha-1). Several other cowpea lines have clear potential for release as new high-yielding varieties with early maturity and high seed quality for farmers in Egypt. Seeds of selected lines are available from Kafrelsheikh University. This shows that mutation breeding and pedigree selection methods are among the most promising breeding methods for cowpea improvement.

A valine residue deletion in ZmSig2A, a sigma factor, accounts for a revertible leaf-color mutation in maize

Abstract: A nuclear-encoded sigma (σ) factor is essential for the transcriptional regulation of plant chloroplast-encoded genes. Five putative maize σ factors have been identified by database searches, but their functions are unknown. We report a maize leaf color mutant etiolated/albino leaf 1 (eal1) that was derived from space mutation breeding. The eal1 mutant displays etiolated or albino leaves that then gradually turn to normal green at the seedling stage. The changes in eal1 leaf color are associated with changes in photosynthetic pigment content and chloroplast development. Map-based cloning revealed that a single amino-acid deletion changing Val480-Val481-Val482 to Val480-Val481, in the C-terminal domain σ4 of the putative σ factor ZmSig2A, is responsible for the eal1 mutation. In comparison with the expression level of the wild-type (WT) allele ZmSig2A+ in WT plants, much higher expression of the mutant allele ZmSig2AΔV in eal1 plants was detected before the eal1 plants turned to normal green. ZmSig2A shows the highest similarity to rice OsSig2A and Arabidopsis SIG2. Ectopic expression of ZmSig2A+ or ZmSig2AΔV driven by the cauliflower mosaic virus 35S promoter rescued the pale green leaf of the sig2 mutant, but ectopic expression of ZmSig2AΔV driven by the SIG2 promoter did not. We propose that the Val deletion generated a new weak allele of ZmSig2A that cannot completely abolish the ZmSig2A function. Some genes involved in chloroplast development and photosynthesis-associated nuclear genes showed significant expression differences between eal1 and WT plants. We conclude that ZmSig2A encoding a σ factor is essential for maize chloroplast development. The eal1 mutant with a weak allele of ZmSig2A represents a valuable genetic resource for investigating the regulation of ZmSig2A-mediated chloroplast development in maize. The eal1 mutation may be useful as a marker for early identification and elimination of false hybrids or transgene transmission in the application of genetic male sterility to commercial hybrid seed production.

Determining the optimum gamma irradiation dose for developing novel maize genotypes

Abstract: Mutation breeding has been used successfully worldwide to generate crop varieties with various traits, including pest resistance. Before any mutation breeding is initiated, radio-sensitivity tests ...

Induced variability and assessment of mutagenic effectiveness and efficiency in sorghum genotypes [Sorghum bicolor (L.) Moench].

Abstract: Purpose Sorghum is an important cereal crop, cultivated for food, fodder and biofuel. Mutation breeding techniques are used to create genetic variability for qualitative and quantitative traits in crop plants. The purpose of this study was to create induced variability and estimate mutagenic effectiveness and efficiency of physical and chemical mutagens in two sorghum cultivars. Materials and methods Gamma rays (100, 200, 300 and 400 Gy, Co60 source, Bhabha Atomic Research Center, Mumbai, India), ethyl methane sulphonate (0.1%, 0.2%, 0.3%, and 0.4% EMS, Sigma-Aldrich, India) and their combinations were used to mutagenize 296B (rainy season) and Parbhani moti (post-rainy) cultivars. Morphological and yield traits were analyzed for enhanced variability in qualitative and quantitative traits across M2 and M3 generations. Results Based on the mutagenic sensitivity, lethal dose at 50% survivability (LD50) for both the genotypes was found to be 269-281 Gy in case of gamma rays and 0.32-0.33% for EMS. Based on reduced germination and survivability, mutagenic sensitivity was dose dependent and genotype independent. High frequency of chlorophyll mutations (albino, xantha, viridis, variegated and chlorine) were linearly correlated with dose in both the genotypes. Among the favorable mutants, dwarf and brown midrib were isolated from Parbhani moti population, which could be used in the cross breeding programs. A combined treatment, 100Gy +0.1% EMS showed high mutagenic effectiveness and efficiency. Enhanced genetic variation for quantitative traits as measured by wide range values and coefficient of variation was attributed to the effect of physical and chemical mutagens. Early flowering, high grain yield (24-49% increase over control) were identified in M2 and validated in M3 generation in both genotypes. Conclusions This study has revealed wide genetic variability and better effectiveness and efficiency of the physical (300 Gy) and chemical mutagens (0.2%) and their combination (200 Gy+ 0.2%) across two sorghum genotypes. Significant correlations identified between quantitative traits will help in better selection in the segregating generations.

Early flowering, good grain quality mutants through gamma rays and EMS for enhancing per day productivity in rice ( Oryza sativa L.).

Abstract: PURPOSE Rice is the prominent crop of world and it frames the important component of food chain. But, long duration of highly preferred rice varieties requires more water and are prone to biotic and abiotic stresses. Short duration rice varieties serve as a promising alternative in such cases. Hence, the present study was taken up to induce early flowering mutants in popular rice variety Bapatla 2231 (BPT 2231), a long duration medium slender rice variety using gamma rays and Ethyl Methane Sulfonate (EMS) treatment combinations. MATERIALS AND METHODS Five hundred well filled and viable grains of optimum moisture content (12%) were subjected to irradiation with 8 doses comprising 100 Gy, 200 Gy, 300 Gy, 350 Gy, 400 Gy, 450 Gy, 500 Gy and 600 Gy treatments. For combination treatments, the irradiated seeds of gamma rays were further treated with 30 mM concentrations of Ethyl Methane sulfonate (EMS). The irradiated population were raised up to M3 generation for the identification of early maturing mutants. The isolated early maturing mutants in M3 generation were characterized for phenotypic, biochemical and grain qualities. RESULTS The frequency of desirable early maturing mutants was observed more in combination treatments than the individual treatments of gamma rays. A total of 34 early maturing mutants (M) with variation for grain quality and biochemical traits were obtained. The mutants M-1, M-3, M-5, M-32, and M-34 were high yielding with fine grain type. The mutants M-4, M-17, M-18, M-19, M-20, and M-26 were high yielding with bold grain type. The mutants M-22, M-23, M-24, M-27, and M-28 were high in nutrient content. The early mutants matured 40-50 days earlier than control. CONCLUSION In this variety, the combination treatments showed higher mutation frequency, mutagenic effectiveness and efficiency than the gamma rays in induction of both chlorophyll and viable mutants. This study revealed that the combination treatments were more effective in inducing early mutants than the gamma ray treatments alone. The early maturing mutants can be released as variety after evaluation under different yield trials and the mutants with increased nutrient content and varied grain quality can be utilized in hybridization/heterosis breeding programs for rice crop improvement.

Outcome of UV-B exposure and induction of some chlorophyll phenodeviants in two important hepatoprotective ethnomedicinal wild plants

Abstract: Present piece of work has been performed with an aim to engender genetic variations in Andrographis paniculata (Burm f.) Nees and Phyllanthus niruri L. since both plants own low or very poor genetic variations due to wild nature. A. paniculata and P. niruri both are magnificent hepatoprotective wild medicinal plants which have been used since ancient times as an ethnomedicine to cure several common and chronic ailments with the high competence and less side effects. UV-B radiations induce mutations because they are absorbed by major biomolecule predominantly by proteins and nucleic acids chiefly DNA. Owing to enormous potential as herbal medicines, both plants i.e. Andrographis and Phyllanthus have been selected for mutation breeding experiments using Ultraviolet-B radiations (UV-B) as a mutagen. When germinating seedlings of A. paniculata and P. niruri were reached up to 1–3 cm, they were treated with UV-B radiations for 0 min, 10 min, 20 min and 30 min with a recovery period of one hour at room temperature and were planted in earthen pots in triplicates. During observations, significant variations in growth and pigment content have been observed in both plants (A. paniculata and P. niruri) in a dose based manner. A wide spectrum of chlorophyll phenodeviants (chlorophyll deficient mutants) in M2 generation such as xantha, xanthoviridis, alboviridis, virscent and chlorina mutants in A. paniculata and variegated plant, xanthoviridis, xantha and albino mutants in P. niruri have also been observed. Out of all the chlorophyll mutants obtained, few were lethal hence not survived later, while rest were survived till different stages of development. On the basis of occurrence of chlorophyll phenodeviants in Andrographis and Phyllanthus, mutagenic effectiveness and efficiency of different doses of UV-B rays have been indexed. The practice of indexing of effectiveness and efficiency of any mutagen is being used for the successful execution of mutation breeding programs to find the optimum dose that may facilitate induction of a multitude of other lucrative mutations.

Gamma-rays and EMS induced resistance to mungbean yellow mosaic India virus in mungbean [Vigna radiata (L.) R. Wilczek] and its validation using linked molecular markers.

Abstract: Purpose Mungbean yellow mosaic India virus (MYMIV) is a serious constraint in the mungbean which is a potential source of easily digestible high-quality proteins, fibers, minerals, and vitamins in Asian countries. Developing resistant cultivars is the most cost-effective, eco-friendly, and sustainable approach to protect mungbean from MYMIV damage. Mutation breeding provides a quick and cost-effective way of developing resistance as lack of genetic variability is the biggest bottleneck for other traditional breeding tools. Materials and methods Outstanding but MYMIV-sensitive varieties of mungbean viz., MH 2-15 and MH 318 were mutagenized through various individual and combined doses of gamma-rays and Ethyl methanesulfonate (EMS) and evaluated in M2 and M3 generations for the appearance of resistance reactions. This was subsequently validated through marker-assisted genotyping using previously reported Yellow Mosaic Disease (YMD) linked markers. Results The phenotyping in M3 generation yielded 64 MYMIV resistant mutants whereas, marker-assisted genotyping identified the 22 mutants with true resistance. Markers YR4, CYR1, and CEDG180 were found associated with MYMIV resistance whereas, DMB-SSR158 did not show any amplification. Among identified resistant mutants, ten lines exhibited at par and two revealed a little higher seed yield over controls. Conclusions: The mutagenesis created significant variability in MYMIV resistance as well as seed yield per plant. YR4, CYR1, and CEDG180 are found to be linked with the MYMIV loci in the mungbean and could be utilized for MYMIV resistance breeding. Mutant M-37 from MH 2-15 and M-104 from MH 318 exhibited more seed yield along with MYMIV resistance which upon further validation can be released as a variety. The induced mutagenesis integrated with powerful emerging molecular and next-generation sequencing (NGS) tools would be highly helpful in breeding mungbean for durable resistance against threatening MYMIV.

Studies on Gamma Rays Induced Cyto-Morphological Variations and Procurement of Some Induced Novel Mutants in Kalmegh [Andrographis paniculata (Burm. f.) Nees]

Abstract: In order to increase genetic variability for improvement and better adaptation in the present days' changing climate, present investigation has been done in Andrographis paniculata (Burm. f.) Nees (also known as Kalmegh in Ayurveda), an ethno-medicinal wild plant with highly immune-boosting properties. For the purpose, healthy, dry and homogeneous seeds were treated with four doses of gamma rays from Co-60 sources at the dose rate of 1.55 Gy per second; selected on the basis of LD50 i.e. 25 Gy (gray), 50, 100 and 200 Gy along with one set of non-irradiated seeds and sown in triplicates in a completely randomized block design (CRBD). Higher doses were detrimental hence not suggested for further mutation breeding experiments. However, lower doses (viz. 25 and 50 Gy) of gamma rays were stimulatory; had induced some significant (p > 0.5) variations in several lucrative traits of Andrographis paniculata (Kalmegh) viz. Plant height, leaf area, number of branches per plant, length of internodes, fruit length, and seeds per fruit. Some induced novel mutants (viz. bushy, dark green leaves mutant and tricotyledonous leaf mutant in M2 generation) have also been procured. This is, to the best of our knowledge, first report on induction of tricotyledonous leaf mutant in A. paniculata using gamma rays.

Determination of the effective radiation dose for mutation breeding in purple carrot (Daucus carota L.) and possible variations formed.

Abstract: Plant breeding allows altering the genetic structure of plants to meet human needs. The use of radiation technology for inducing mutations and -thereby- new phenotypic variants has become increasingly common as a tool for developing new crops. The aim of this study was to determine the effective gamma irradiation dose for inducing mutations in purple carrot. Increasing gamma radiation doses [0, 50, 100, 200, 300, 400, 500, and 600 Gy] were applied to purple carrot seeds. The irradiated seeds were sown in pots and the emergence and survival rates of the seedlings were analyzed. Considering plant emergence (%) as a response variable, the LD50 dose was 387.5 Gy. Analysis of root length, root width (shoulder diameter) and plant height in control (0 Gy) and irradiated plants (50–600 Gy) revealed an inverse association between these morphological traits and radiation dose. SRAP and ISSR markers were used to identify DNA polymorphisms in irradiated and control plants. The range of amplicons per primer set revealed by ISSR and SRAP markers was 4–10 and 2–13, respectively. In the ISSR analysis of the irradiated carrots (for the 8 doses used), we obtained range values for the average Nei’s gene diversity, Shannon’s information index, and polymorphism information content (PIC) of 0.13–0.25, 0.20–0.35, and 1.39–1.67, respectively, whereas in the SRAP analysis, the range values for these parameters were 0.15–0.25, 0.23–0.37, and 0.43–0.58, respectively. Cluster analysis revealed three main groups; (a) non-irradiated (control) plants, (b) plants from the 600 Gy dose, and (c) a third group with two subgroups: one with individuals from the lowest irradiation doses (50–200 Gy) and a second group with individuals from the highest irradiation doses (300–500 Gy). This is the first report on determining effective mutagen doses and genetic characterization of induced mutagenesis via gamma irradiation in purple carrot. ISSR and SRAP markers were successful in detecting variations among different levels of mutagen doses.

Studies on mutagenic sensitivity of seeds of pummelo (Citrus maxima Merr.)

Abstract: Mutation breeding is a key method of generating a large number of heritable variations. Effective dose (LD50) needs to be standardized for inducing sufficient variation in a crop. In the present study, seeds were irradiated with different doses of Gamma rays and found that 66.94 Gy could suppress germination close to 50 percent (LD50) in pummelo. This 60 Gy gamma dose can effectively be used for raising the mutant populations to identify a desirable mutation in pummelo.

Improving seed yield and protein content of common beans (phaseolus vulgaris l.) through mutation breeding

Abstract: The common bean is a staple food and source of protein in many African countries. To date, genetic improvement of common bean in Egypt lags behind compared with other legume crops which mainly due to limited genetic resources. This study was carried out in three consecutive growing seasons (2016, 2017 and 2018) at Sakha Experimental Station, Horticulture Research Institute (HRI) at Kafr El-Sheikh governorate, Egypt. The study aimed to increase genetic variations in Nebraska bean cultivar through mutation breeding for selection of novel bean lines with higher seed yield and protein content. The experiment was arranged in a randomized complete blocks design (RCBD) with three replications. Seeds of Nebraska bean cultivar which widely grown in Egypt were treated with gamma rays three different doses (10, 20, and 30 kr), and selection was made at M2 and M3 generations for desired traits related to vegetative growth parameters, pod characteristics and seed yield components. These traits were significantly affected by irradiation treatments in M1 generation, and changes in plant height, number of branches per plant, number of pods and seeds per plant were generally favorable in M2 generation than control (un-irradiated plants). Analysis of variance in M3 generation showed a highly significant difference among M3 lines and control for vegetative and agronomical traits except for the traits of number of days to first flower and number of seeds per pod. Among 25 M3 lines, ten mutant lines were produced higher seed yield (4.3 to 108.6%) and protein content (0.08 to 2.79%) along with heavier seeds (3.5 to 35%) than original parent Nebraska cultivar. Hence, these lines should be evaluated for the desired traits and wide adaptability to biotic and abiotic stress to release new improved varieties in Egypt.