Ethyl Methanesulfonate Concentration Research Articles

PRODUCTION MUTANT LINES OF TURNIP RAPE (BRASSICA RAPA) AND ITS INTERSPECIFIC HYBRIDS IN THE ISOLATED MICROSPORE CULTURE

This article presents the results of research on the production of mutant doubled haploids of turnip rape (Brassica rapa L.ssp.Oleifera), as well as its interspecies hybrids with rapeseed (Brassica napus). An  immature embryo culture  was used to increase the productivity of inter-specific hybridization. Studies have obtained interspecific hybrids of turnip rape with rapeseed. The largest number of plants was observed in hybrids where the mother plant was rapeseed. Furthermore, chemical mutagenesis was used to increase the genetic diversity of studied materials. Ethyl methanesulfonate (EMS) has been applied to the treatment of embryos produced in the isolated microspores culture  of turnip rape and its interspecies hybrids. Results of embryo regeneration against mutagen on a nutrient medium B5 with gibberellic acid showed that the concentration of 8 mM EMS stimulated the regeneration process in turnip rape cultivars (Zolotistaya - 72% and Yantarnaya - 88%) and showed superior results compared to a low concentration (4 mM). For hybrids compared to high concentration (8mM), a low concentration of 4mM EMS showed superior results. New mutant lines have been obtained as the starting material for selection in the creation of new domestic canola cultivars.

Development and characterization of ethyl methane sulfonate (EMS) induced mutant population of grass pea with yield, quality and forage traits

Grass pea (Lathyrus sativus L.) is an important protein-rich annual legume crop that can survive harsh environment conditions. Genetic improvement of grass pea is highly important for expanding its production, and mutation breeding can be a valuable tool in creating additional genetic variability. In the present study, we treated totally 3000 seeds of widely cultivated grass pea cultivar, Gürbüz-2001, with two different concentrations (0.5% and 1%) of mutagen EMS for 4 hours. The germination rates were calculated as 71.6% (1074 plants) and 42.4% (636 plants) for 0.5% and 1% EMS concentrations in M1 generation, respectively. We identified several mutant phenotypes such as seedling growth defects, abnormal branching, chlorophyll mutants, absence of flowering, and sterility in M1 and M2 populations. A total of 40 mutant plants which were superior than control variety in terms of vegetative components from M2 population were selected for agronomic, quality and forage traits analysis. Plant height of mutant lines ranged from 48 (GMP29) to 164 cm (GMP4), with an average value of 109.7 cm. One of the most interesting features of some mutant lines was the significant increase in 1000-seed weight over the control value, contributing to higher yield. The seed yield ranged from 7.19 to 87.18 g, and the lines GPM6 and GMP11 showed superior values compared to the control (13.6 g), indicating the positive effect of mutation in obtaining desired traits. There were promising mutants that showed higher levels of protein content among mutated lines. The β-ODAP (β-N-Oxalyl-l-α,β-diaminopropionic acid) content of grass pea seeds ranged from 0.139% to 7.39% in selected M2 population, while the control cultivar was 0.523%. GPM6 and GPM11 were important for forage breeding with their high crude protein content, low ADF (acid detergent fiber) and NDF (neutral detergent fiber) ratios, as well as high seed and biological yield. The beneficial traits of these mutants obtained in this study should be followed to integrate as genetic resources for use in grass pea improvement programs.

Increasing the diversity of marigold (Tagetes sp.) by acute and chronic chemical induced mutation of EMS (Ethyl Methane Sulfonate)

Abstract. Lenawaty DY, Sukma D, Syukur M, Suprapta DN, Nurcholis W, Aisyah SI. 2022. Increasing the diversity of marigold (Tagetes sp.) by acute and chronic chemical induced mutation of EMS (Ethyl Methane Sulfonate). Biodiversitas 23: 1399-1407. Increasing the diversity of marigold (Tagetes sp.) can be carried out through chemical induction mutations with Ethyl Methane Sulfonate (EMS).This study aims to obtain the LC50value, determine the sensitivity level of plants, and obtain plant diversity of Tagetes sp.with EMS both acutely and chronically. The genetic material used in this study was derived from T. erecta genotype MG04 and T. patula genotype MG21. The acute mutation technique was performed by soaking the seeds in EMS concentration for 4 hours under 10 different concentration levels (0%, 0.2%, 0.4%, 0.6%, 0.8%, 1.0%, 1.5%, 2.0%, and 3.0%). The chronic mutation technique was carried out by diluting the concentration of EMS LC501/10x, 1/100x, 1/1000x and immersion time of 6 hours, 24 hours and 48 hours. The study used a Completely Randomized Block Design. Observationswere made on the percentage of surviving plants, quantitative and qualitative characters. The data obtained were analyzed using Boxplot and ANOVA. The results showed that the LC50 value of T. erecta is 0.82% and LC50of T. patula is 1.87%, which means that the sensitivity level of T. erecta to EMS treatment is quite higher than T. patula. The acute application method shows a relatively low mutation rate and limited diversity of flower types. The chronic application method produced various flower shapes and whiter flower color in T. erectaand T. patula resulting in a redder flower color than the acute application method.

ASSESSMENT OF SEED GERMINATION IN BLACKGRAM (VIGNA MUNGO L. HEPPER) UNDER EMS INDUCED MUTAGENESIS

The present investigation was carried out to estimate the mutagenic sensitivity of blackgram accession IC 436524 with EMS (Ethyl methane sulfonate). The seeds of IC436524 were treated with four different concentrations of EMS viz., 0.2%, 0.3%, 0.4% and 0.5%. Thirty seeds of each concentration were placed in Petri plates in three replication along with untreated seeds for germination test, shoot length(cm) and root length(cm). The germination was observed on the 5th day after treatment while shoot and root length of seedling was recorded on the 10 day. The results showed that the germination% was recorded highest (86.67%) at 0.2% of EMS followed by (80.00%) at 0.3% of EMS and lowest seed germination% was recorded at 0.4 and 0.5% of EMS. The reduction percentage of seed germination with control (untreated seeds) at 0.2% of EMS was 6.66% followed by 13.33% at 0.3% of EMS, 53.33% at 0.4% of EMS and 80% at 0.5% of EMS respectively. The results also showed that the highest shoot and root length (cm) was recorded at 0.2% of EMS (8.60cm and 5.50cm) followed by at 0.3% of EMS (6.50cm and 3.83cm). Thus 0.4 and 0.5% dose of EMS seemed very effective in reducing percent seed germination in black gram. The results excerted that the blackgram accession IC436524 could be germinated up to 0.3% EMS concentration to create variability in blackgram which are further useful for crop improvement programs.

Directed evolution of Chlorella sp. HS2 towards enhanced lipid accumulation by ethyl methanesulfonate mutagenesis in conjunction with fluorescence-activated cell sorting based screening

The present study aimed to improve the lipid content and lipid productivity of Chlorella sp. HS2 by exposing them to ethyl methanesulfonate (EMS) to generate a mutant and followed with fluorescent assorted cell sorting (FACS) for the selection of mutant cells having desirable phenotypes. The microalgae were subjected under 100 μ mol mL−1 concentration of EMS for 30 min, followed by intracellular lipid staining with boron-dipyrromethene (BODIPY) fluorophore for FACS based selection. The biomass growth, lipid content, lipid productivity, and fatty acid composition of the selected mutagenized strain (SMS, TE+F) of Chlorella sp. HS2 were compared against those of FACS selected strain without mutagenesis (FSS, TF) and parent wild strain (PWS, TC), under two different carbon supplementation schemes (only CO2 supply and optimized carbon supply). The highest lipid content and productivity of 35.6% and 248.7 mg L-1 d-1, respectively were observed in the SMS grown under optimized carbon supplementation (1% CO2 (v/v) and 0.5 g L-1 NaHCO3), which was a substantial improvement over those of FSS (26.6% and 193.4 mg L-1 d-1) and PWS (24.7% and 153.6 mg L-1 d-1). The fatty acid methyl ester (FAME) profile of the lipids derived from SMS showed a marked increase in the proportion of saturated fatty acids (SFA) and mono-unsaturated fatty acids (MUFA) (76.7%) over those of FSS (70.2%) and PWS (69.6%). Lastly, fuel properties were found to be appropriate for biodiesel production, as they corroborate with international fuel standards. These results indicate that directed evolution using EMS mutagenesis with FACS is a powerful tool to achieve microalgal strain improvement for biofuel production.

ANALYSIS OF COURTSHIP BEHAVIOR AS A PARAMETER OF TOXICITY OF ETHYL METHANESULFONATE (EMS) IN DROSOPHILA MELANOGASTER

An anti-cancerous drug and a monofunctional alkylating agent, Ethyl methanesulfonate (EMS), was tested for its toxic effects by using courtship behavioral elements as the parameter in Drosophila melanogaster. EMS concentrations ranging from 0.5 to 15mM were employed by larval feeding technique to analyze the effect. When the different combinations of crosses were performed, the results have revealed that there were no qualitative differences between the control and treated groups. However, quantitatively, it has shown that male courtship elements such as orientation, tapping, wing vibration and licking were significantly prolonged compared to controls. Similar results were observed in case of female rejection behavior elements namely: ignoring, extruding and decamping. On par with this, even the courtship latency and copulation latency were significantly increased and contrary to these, copulation duration was significantly decreased, thus suggesting that the treated males were less vigor; treated females were less receptive on one hand and on the other lower reproductive fitness. Drastic effects of EMS on mating behavior prove that this can be utilized as a parameter for assessing the toxic effects of chemicals.

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

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.

Physiological and biochemical responses of Kinnow mandarin (Citrus nobilis × Citrus deliciosa) to EMS induced mutagenesis

The present study was carried out to enhance the genetic variability in Kinnow mandarin (Citrus nobilis Loureiro Physiological and biochemical responses of Kinnow mandarin (Citrus nobilis × Citrus deliciosa) to EMS induced mutagenesis Citrus deliciosa Tenora) using ethyl methanesulfonate (EMS) during 2016-17 and 2017-18. Plants created with different doses of EMS concentration, viz. 0.05%, 0.1%, 0.2% and 0.5% were examined for the various physiological and biochemical alterations in putative mutants aged six years and compared with the wild type (WT). A dose-dependent decrease in leaf area was recorded in the mutants with increasing doses of EMS concentration and as compared to WT, maximum reduction of 135.52 and 104.92% was witnessed in the mutants E-19 and E-17 developed from 0.5% EMS, whereas the leaves were more succulent in E-16. Chlorophyll a content was higher than chlorophyll b in the mutated population. As compared to WT, maximum decrease in chlorophyll a, total chlorophyll was noticed in the mutants E-19 generated from 0.5% EMS. Contrary to the physiological alterations, the total phenol, proline and total protein content in the mutants were upregulated by nearly two fold in the mutants developed beyond 0.1% EMS. Significant increase was also noticed in the antioxidant enzymes SOD, CAT, POX and GR activity in the mutants E-16 to E-20 induced from 0.5% EMS. The variation in traits of interest observed in the present study such as better leaf succulency and upregulated antioxidant enzyme activity in the mutant E-16 (0.5% EMS) suggests that the developed mutant could be utilised in Kinnow improvement for abiotic stress tolerance.

Investigation effect of ethyl methane sulfonate (EMS) on some of morphophysiological and phytochemical traits of fenugreek (Trigonella foenum-graecum L.)

Fenugreek (Trigonella foenum-graecum L.) is a member of the Fabaceae family which is an annual, dicotyledonous and self-pollinated plant. Since genetic diversity is the basis of plant breeding programs, it is important to create genetic mutations in order to increase diversity. To induce mutation, this experiment was conducted by applying four levels (0.1, 0.2, 0.3, and 0.4 %) of ethyl methane sulfonate (EMS) on fenugreek seeds. The treated seeds were planted in the research greenhouse on basis of randomized complete block design with three replications. The results showed that different EMS concentrations had significant effect on all the studied characteristics. The lowest values of the studied traits were observed in the control. Although the highest values of most morpho-physiological traits were obtained in 0.2 % of EMS, but the highest amount of phytochemical compounds were related higher concentrations of EMS. The content of trigonelline, diosgenin, nicotinic acid, and mucilage of grain were significantly increased with increasing EMS concentration. The highest and lowest contents of grains trigonelline were obtained in 0.3 % of EMS and control treatment, respectively. The nicotinic acid content had a positive and significant correlation with trigonelline, diosgenin and mucilage content. Also, the diosgenin content of grains had a positive significant correlation with mucilage content. In general, the application of sufficient concentrations of EMS mutants could be used as an effective tool to increase grain yield and secondary metabolites in breeding programs of fenugreek.

Development of EMS-induced Mutagenized Groundnut Population and Discovery of Point Mutations in the <i>ahFAD2</i> and <i>Ara h 1</i> Genes by TILLING

Reducing allergenicity and increasing oleic content are important goals in groundnut breeding studies. Ara h 1 is a major allergen gene and Delta(12)-fatty-acid desaturase (FAD2) is responsible for converting oleic into linoleic acid. These genes have homoeologues with one copy in each subgenome, identified as Ara h 1.01, Ara h 1.02, ahFAD2A and ahFAD2B in tetraploid groundnut. To alter functional properties of these genes we have generated an Ethyl Methane Sulfonate (EMS) induced mutant population to be used in Targeting Induced Local Lesions in Genomes (TILLING) approach. Seeds were exposed to two EMS concentrations and the germination rates were calculated as 90.1% (1353 plants) for 0.4% and 60.4% (906 plants) for 1.2% EMS concentrations in the M1 generation. Among the 1541 M2 mutants, 768 were analyzed by TILLING using four homoeologous genes. Two heterozygous mutations were identified in the ahFAD2B and ahFAD2A gene regions from 1.2% and 0.4% EMS-treated populations, respectively. The mutation in ahFAD2B resulted in an amino acid change, which was serine to threonine predicted to be tolerated according to SIFT analysis. The other mutation causing amino acid change, glycine to aspartic acid was predicted to affect protein function in ahFAD2A. No mutations were detected in Ara h 1.01 and Ara h 1.02 for both EMS-treatments after sequencing. We estimated the overall mutation rate to be 1 mutation every 2139 kb. The mutation frequencies were also 1/317 kb for ahFAD2A in 0.4% EMS and 1/466 kb for ahFAD2B in 1.2% EMS treatments. The results demonstrated that TILLING is a powerful tool to interfere with gene function in crops and the mutagenized population developed in this study can be used as an efficient reverse genetics tool for groundnut improvement and functional genomics.

Novel genetic variability in sesame induced via ethyl methane sulfonate

ABSTRACT Sesame (Sesamum indicum L.) is an ancient oilseed crop with important agronomic, therapeutic, and industrial properties. However, in Morocco, the existing genetic diversity is quite limited. Thus, chemical mutagenesis, using ethyl methane sulfonate (EMS), was applied to induce novel genetic variability. Seeds of two sesame genotypes were treated using two EMS concentrations, 0.5% and 1%. The treated and untreated seeds (control) were planted according to a completely randomized design. Some mutant plants (M1) were identified and characterized for growth habit, root length, number of carpels per capsule, number of capsules per leaf axil, seed yield per plant, and seed color. Specifically, mutants with a tetra-carpellate capsule, three capsules per leaf axil, determinate growth, diverse seed colors, and a highly developed root system were found to be promising and useful for sesame breeding program aimed at developing productive and high-quality cultivars, particularly for stressful environments.

Effects of time and concentration of ethyl methane sulfonate (EMS) on chia (Salvia hispanica L.)

Effects of time and concentration of ethyl methane sulfonate (EMS) on chia (<i>Salvia hispanica</i> L.)

The Effect of Ethyl Methane Sulfonate (EMS) on The In Vitro Shoot Regeneration of Vetiver (Vetiveria zizanioides [L.] Nash)

Production of vetiver in Indonesia is low, and its oil quality is not in line with market demand due to the low quality of plant raw materials. In vitro mutagenesis using ethyl methanesulfonate (EMS) has the potential to produce superior vetiver variants. This study aimed to determine the effect of EMS on the growth and shoot formation of in vitro vetiver plants. In vitro mutagenesis was performed by culturing in vitro shoots on MS medium containing EMS with various concentrations (0.02; 0.04; 0.06; 0.08; 0.1%) for 4 weeks. The results showed that the addition of 0.02-0.1% EMS to the medium affected on in vitro shoot growth and the ability to form new shoots. EMS increased the percentage of dead explants and decreased the average number of shoots, the number of shoots formed per explant, and the shoot fresh weight. The higher the EMS concentration in the medium, the smaller the percentage of survived explants and the lower ability to form new shoots with the average number of shoots formed less. The percentage of survived explants on the medium without the addition of EMS and on the medium with the addition of EMS with low concentrations of 0.02 and 0.04% reached 100% with an average number of shoots formed ranging from 8.5-10 shoots/explant. Meanwhile, the percentage of survived explants on medium with the addition of EMS with a high concentration of 0.08-0.1% was only 12.5-25% with an average number of shoots formed was less than 2 shoots/explant. Keywords: EMS, in vitro shoots, Vetiveria zizanioides .

EMS‐induced mutations in common vetch (Vicia sativa L.) and two mutants without anthocyanin accumulation showing increased cold tolerance

AbstractCommon vetch (Vicia sativa L.) is a leguminous plant used as forage and cover crops. A protocol of ethyl methanesulfonate (EMS)‐induced mutations was established in the present study, based on analysis of seed germination and emergence rate, seedling growth and abnormal plant percentage as affected by combinations of EMS concentration and treatment time. Treatment of seeds with 0.1% EMS for 9 and 12 hr or 0.5% EMS for 6 and 9 hr led to 43%–66% normal plants, which was suggested to be used for EMS‐induced mutations in common vetch. The wild‐type common vetch showed red with accumulation of anthocyanins in winter, and two lines showing separation of anthocyanin accumulation (labeled as A+) and without anthocyanin accumulation (labeled as A–) plants were selected from M2 plants induced by EMS in the field. The A– plants showing green color had higher plant height and fresh shoot weight than the A+ plants during wintering, with lower ion leakage after freezing treatment, indicating that the green mutants (A–) had increased cold tolerance. Compared to A+ plants, the green mutants had significantly increased ascorbate (AsA) level in response to low temperature treatment. The results suggest that the increased AsA level under low temperature was associated with the increased cold tolerance in the green mutants.

Promoting the accumulation of scopolamine and hyoscyamine in Hyoscyamus niger L. through EMS based mutagenesis.

The overexploitation of medicinal plants is depleting gene pool at an alarming rate. In this scenario inducing the genetic variability through targeted mutations could be beneficial in generating varieties with increased content of active compounds. The present study aimed to develop a reproducible protocol for in vitro multiplication and mutagenesis of Hyoscyamus niger targeting putrescine N-methyltransferase (PMT) and 6β-hydroxy hyoscyamine (H6H) genes of alkaloid biosynthetic pathway. In vitro raised callus were treated with different concentrations (0.01% - 0.1%) of Ethyl Methane Sulfonate (EMS). Emerging multiple shoots and roots were obtained on the MS media supplemented with cytokinins and auxins. Significant effects on morphological characteristics were observed following exposure to different concentrations of EMS. EMS at a concentration of 0.03% was seen to be effective in enhancing the average shoot and root number from 14.5±0.30 to 22.2 ±0.77 and 7.2±0.12 to 8.8±0.72, respectively. The lethal dose (LD50) dose was calculated at 0.08% EMS. The results depicted that EMS has an intense effect on PMT and H6H gene expression and metabolite accumulation. The transcripts of PMT and H6H were significantly upregulated at 0.03–0.05% EMS compared to control. EMS treated explants showed increased accumulation of scopolamine (0.639 μg/g) and hyoscyamine (0.0344μg/g) compared to untreated.

Exposing Seeds of Galtonia candicans to Ethyl Methanesulfonate Reduced Inflorescence Height, Lodging, and Fertility

Cape hyacinth (Galtonia candicans) is a geophytic herbaceous perennial from South Africa. It produces large inflorescences of pendulous white flowers during mid to late summer, followed by capsules filled with copious amounts of seed. The species has potential as a low-water-use landscape plant, but lodging and excessive seed production, which pose a risk of escape or invasion, are issues that should be addressed before marketing. Ethyl methanesulfonate (EMS) is a chemical mutagen known to induce usable mutations including dwarfing and sterility. We exposed seeds of cape hyacinth to increasing concentrations of EMS (0%, 0.2%, 0.4%, 0.6%, 0.8%, and 1%). Increased concentrations of EMS resulted in a linear decrease in seed germination when not exposed to a presoak treatment in water before exposure to EMS. No seedlings survived or were viable to field plant at 0.6%, 0.8%, or 1%. Resulting plants were field planted in 2013 and evaluated during 2014 and 2015. In both years, the inflorescence height at first flower, average seed number per capsule, and percent lodging were reduced in EMS-treated plants compared with controls. In 2015, pollen staining was evaluated and was reduced from 83% in control to less than 3% in the 0.4% treatment. Our study demonstrated that EMS is a viable option to reduce height and decrease seed set in cape hyacinth.

High frequency of transversion mutations in the rice (Oryza sativa L.) mutant population produced by diepoxybutane mutagenesis

Discovering new genetic mutations is vital for expanding genetic resources for exploring the functions and breeding applications of genes. In this study, we created new mutant populations of rice and evaluated its effectiveness by using a chemical mutagen diepoxybutane (DEB) with the expectation of manageably causing small-scale deletions that induce knock-out mutations. Compared with the more common mutagen, ethyl methanesulfonate (EMS), DEB exhibited approximately 160 times the adverse impact on the early growth of rice. At 0.3 mM, which was 1/160 the concentration of EMS in the control population, the heading date in the DEB-mutated population showed dispersion, albeit with a small standard deviation. Therefore, similar to EMS, DEB has been shown to induce DNA mutations. According to the screening of waxy mutants using the Targeting Induced Local Lesions in Genomes (TILLING) method, DEB-mutated populations had nearly similar mutation frequencies to those of EMS-mutated populations. Therefore, we successfully isolated five mutant lines from the DEB-mutated population. Some of these mutants exhibited a low-amylose phenotype, which is applicable to breeding leading to enhanced taste evaluation. To utilize these mutated alleles, we developed co-dominant DNA markers. In this study, EMS induced transition mutations, as previously reported. In contrast, DEB specifically induced transversion mutations rather than small-scale deletions contrary to our initial expectations. These results demonstrate that DEB has a novel point of action for mutation and is useful for expanding genetic resources for crops.

Effect of chemical mutagens on growth of Okra (Abelmoschus esculentus L. Moench)

The purpose of the present study was to evaluate the effects of mutagens i.e. Ethyl methanesulfonate (EMS) and Sodium azide (SA) on the morphological growth of Okra. Seeds of two Okra varieties Subzpari and Pahuja were first pre-soaked in distilled water and then treated with 0.0 (Control), 0.1, 0.2 and 0.3% Sodium azide (SA) and Ethyl methanesulfonate (EMS) separately. The results showed that the germination rate was significantly decreased with increased lethality (%) and mutation frequency (%) in M1 of both varieties with increasing concentrations of EMS and SA against control. The results showed that different concentrations of EMS and SA affected morphological attributes of both okra varieties as the increase of mutagen concentration showed variable results in M1 generation compared with control, but M2 generation showed useful improvement in various traits. Such as the M2 generation of Subs pari from 0.1% SA treatment showed a decrease in flowering time while an increase in the number of fruits per plant, fruit size, seeds yield (g) per plant and 100 seeds weight. Keywords: Ethyl methane sulfonate; Growth; Induced mutagenesis; Okra; Sodium azide http://dx.doi.org/10.19045/bspab.2020.90116

Sterile leucaena becomes a reality?

Keynote paper presented at the International Leucaena Conference, 1‒3 November 2018, Brisbane, Queensland, Australia.A research program to develop sterile leucaena has commenced to enhance red-meat production in additional regions of Australia including Western Australia, Northern Territory and New South Wales, where growing seeded leucaena is not currently permitted or encouraged. In this study we report on the development of methodology using a mutagenizing agent, EMS (ethyl methanesulfonate), to cause mutations in the self-fertile commercial leucaena cultivar, Redlands. Several experiments to determine the optimum rate of EMS have been completed and first generation mutagenized plants (M0) established in the field at Redlands Research Station, Cleveland, Queensland, Australia. An EMS concentration of 0.35% applied to germination paper proved the best method to achieve a target emergence percentage of 50%. To date, 27 of 179 mutagenized M0 seedless plants are considered to be putatively sterile. A further 1,200 M0 plants have been established in the field providing an even greater chance of identifying sterile leucaena plants with the desired forage quality and psyllid-resistance attributes.

A new male sterile line of “Duanye-13” radish (Raphanus sativus L.) produced by ethyl methanesulfonate mutagenesis

Mutations in radish can be induced by exposure to ethyl methanesulfonate (EMS), and this method is suitable for obtaining target traits in new radish materials. We investigated the optimal concentration of EMS for mutagenesis on the radish variety Duan-Ye No. 13 (DY13). Treating DY13 radish seeds with 0.5% EMS led to the development of sterile mutants. During 3 years of continuous hybridization and tracing, male fertility remained stable at 100% sterility. Morphological observations showed that the anthers of these male sterile were wrinkled, curled, and browned; disintegration of the wall structure at the pollen tetrad stage caused eventual pollen abortion. Detection of the molecular marker ORF138-F/R confirmed that the target band could be amplified in this radish line; however, the band was not amplified in the corresponding maintainer line. Together, these results show that male sterile lines were successfully selected from DY13 radishes, demonstrating a new and effective method for breeding male sterile radish lines.