M2 Generation Research Articles

Morphology Performance On Six Black Rice Accessions (Oryza Sativa L.) In M1 Generation Irradiated By Gamma Rays

Increasing the genetic diversity of plants can be done through physical mutations with gamma-ray irradiation. This study aims to determine gamma-ray irradiation on the growth of six M1 generation black rice accessions. This research was conducted in May-October 2019 at the Experimental Rice Fields of the Tasikmalaya Perjuangan University, West Java. The materials used in this study were six accessions from Tasikmalaya (PH, PH2, PH3, PH5, PH7, and PH8) without 0 Gy irradiation and with 200 Gy gamma-ray irradiation of M1 generation. The results showed that gamma-ray irradiation reduced germination characteristics, seedling height, and flowering age. Some of the accessions showed different effects; there were a decrease and an increase in the characters of root length, plant height, number of productive tillers, number of unproductive tillers, and the total number of tillers.

Efficiency of electron beam over gamma rays to induce desirable grain-type mutation in rice (Oryza sativa L.)

Purpose Rice is the predominant crop of Tamil Nadu state, India that occupies about 30% of the total cropped area. However, grain type and quality are the critical traits that determine the market value and domestic consumption rice variety. Most of the households of Tamil Nadu, India prefer to consume medium slender to fine grain type of rice. Hence, the present study was conducted to induce medium slender grain type in popular rice variety ADT 37 (Aduthurai 37), a short bold rice variety using gamma rays (GR) and electron beam (EB) mutagens. Materials and methods Healthy, dried seeds (12.0% moisture content) of ADT 37 rice variety were exposed to various doses of GR (100–500 Gy) and EB (200–600 Gy). The irradiated population were maintained up to M4 generation by plant to progeny row basis to identify stable mutants for grain-type variation. The selected grain-type mutants (medium slender- and slender-type mutants) in M4 generation were characterized for phenotypic and grain quality traits. Results A high frequency of desirable grain-type variation was observed in EB-irradiated population than gamma-irradiated population. A total of 25 grain-type mutants (long slender and medium slender) were obtained in M4 generation of ADT 37 variety. The morphological characterization and cooking quality assessment of the ‘grain-type’ mutants revealed that six out of 25 mutants viz., M-3 (Mutant-3), M-5, M-9, M-10, M-13 and M-15 recorded single plant yield of more than 30 g. There was non-significant variation in yield per plant (g) among the mutants and control (parent) due to key changes in grain type and thousand grain weight. Conclusion EB showed higher mutation frequency, mutagenic effectiveness and efficiency than the GR in inducing both chlorophyll and viable mutants. This study revealed that the percentage contribution of the EB was 2.57 times higher than that of GR in obtaining desirable slender and medium slender grain-type mutants. The grain-type mutants obtained in the present study can be either directly released as variety or used as parents in hybridization program of rice crop improvement.

Estimation of variability, heritability and genetic advance in M3 generation of natural mutant of dolichos bean

An experiment entitled “Estimation of variability, heritability and genetic advance in M3 generation of natural mutant of dolichos bean” was carried out at All India Coordinated Research Project on Vegetable Crops, Mahatma Phule Krishi Vidyapeeth, Rahuri, Dist. Ahmednagar (MS), which was conducted during the year 2018-19 with an objective to assess the genetic variability in M3 generation. The estimates of phenotypic coefficient of variation were higher than that of genotypic coefficient of variation for all the characters under study in M3 generation. High GCV and PCV were observed for vine length, number of racemes per vine, fresh pod width, number of fresh pods per vine, fresh pod yield per vine, fresh pod yield per hectare, fresh seed yield per vine and dry seed yield per vine in M3 generation indicating the higher magnitude of variability and direct selection of these traits may be advantageous in developing desirable dolichos bean genotype. The high heritability along with high genetic advance as percent of mean are exhibited by vine length, days to 50 per cent flowering, number of racemes per vine, pod setting per cent, fresh pod length, fresh pod width, number of fresh pods per vine, fresh pod shelling per cent, fresh pod yield per vine, fresh pod yield per hectare, fresh 100 seed weight, fresh seed yield per vine, dry 100 seed weight and dry seed yield per vine in M3 generation. This indicates presence of additive gene action for the expression of these traits which is fixable for next generation and selection in next population based on this character would be ideal.

Determination of morphological changes using gamma irradiation technology on capsicum specie varieties

Abstract The recent increase in the demand of hot and chili pepper at local, national, and international market has drawn the interest of breeders’ crop improvement researchers globally. Thus, there is a need to enhance its breeding and development to improve the production of hot and chilies pepper in the quest for food quality and security. This study focused on the germination ability and evaluation of chlorophyll mutations in M2 generation of gamma irradiates on pepper cultivars. The materials used in this experiment were obtained from the M1 generation of gamma-irradiated seeds of pepper cultivars (Capsicum frutescens, Capsicum chinense, and Capsicum annuum (hot pepper, bonnet pepper, and bell pepper, respectively)). Seeds of the pepper varieties were subjected to doses of gamma ray (i.e., 0, 100, 150, 200, 250, 300, 350, and 400 Gy) after which they were planted, raised, and harvested in the M2 generation. The parameters evaluated were germination percentage, survival percentage, epicotyl height, and chlorophyll mutations. However, the highest chlorophyll mutation frequency was observed in C. frutescens. This showed that it was more sensitive to gamma irradiation relative to C. chinense and C. annuum. virescenes were the highest chlorophyll mutant observed in all the three cultivars, while the least was Xanthan. The effective dose ranges between 50 and 250 Gy for germination and survival rate for the three cultivars of pepper give higher desirable mutations in the pepper cultivars. While 400 Gy recorded most efficient for chlorophyll mutation.

The effect of gamma radiation on the growth of black rice plants generation m1

Black rice is a food that can be consumed as a functional food ingredient. Black rice has not been widely cultivated while the seeds are not commercially available. Besides, black rice also has several weaknesses such as longevity, high plant habitus and low production. This study aims to test black rice accession using gamma ray irradiation in the M1 generation. The research was conducted in January - April 2020 in the experimental rice fields of Cineam District, Tasikmalaya Regency. The materials used are black rice seeds (PH2, PH3, PH5, PH7 and PH8). The treatments tested were 5 black rice accession (PH2M1, PH3M1, PH5M1, PH7M1 and PH8M1) that was irradiated with gamma rays (200 Gray) and 0 Gray as control. The results showed that irradiated and non-irradiated rice growth gave different responses. All black rice accessions after irradiation experienced stem shortening and decreased number of filled grains. Accession PH 7 showed shorter flag leaves, number of productive tillers and fewer total tillers after irradiation. Meanwhile PH8 showed the number of productive tillers and total tillers were more and the panicles were shorter after irradiation.

Effectiveness and efficiency of electron beam in comparison with gamma rays and ethyl methane sulfonate mutagens in cowpea

Cowpea is the poor man's crop that lacks variability due to its autogamous nature. Induced mutation serves as a potential source in the induction of variability in crops. On the other hand, the effectiveness and efficiency of mutagens will vary between species and even varieties. In the present study, a novel mutagen electron beam was used in cowpea for the first time along with commonly used mutagens gamma rays (physical) and ethyl methanesulfonate (chemical). The biological damages on eight quantitative characters in M1 generation and chlorophyll mutants in M2 generation were recorded. Two popular varieties viz., P 152 and VBN 1 constituted as the biological material of study. The rate of reduction in biological damage on quantitative characters was directly proportional to the dose of mutagen irrespective of the varieties and mutagens used. Physical mutagens showed the highest biological damage (EB- 37.5% and G- 37.3% overall reduction from control) than chemical mutagen (EMS- 30.4%). Comparing the physical mutagens at similar doses, 200 Gy or 300 Gy of electron beam showed more biological damage than 200 Gy or 300 Gy of gamma rays. Eleven different types of chlorophyll mutants were identified in the M2 generation. Xantha is the most occurred chlorophyll mutants (44.44%), while aurea and yellow viridis have least occurred mutants. Chemical mutagen (EMS) is considered to be the most effective (6.47%) and efficient mutagen (27.09%) based on the chlorophyll mutants and it was followed by an electron beam and gamma rays. Among the physical mutagens, electron beam showed the highest biological damage (37.50% overall reduction from control) and higher effectiveness and efficiency (3.80% and 23.38%) compared to gamma rays (1.87% and 13.38%). Hence, the electron beam can also be used as an effective mutagen in creating variation in cowpea and other crops as it is highly effective, cost less and safe mutagen.

Effect of Gamma Irradiation on Chlorophyll Content in the Cowpea (Vigna unguiculata (L.) Walp)

The experiment was conducted to study the mutagenic effect of gamma rays on chlorophyll content at three different physiological stages in the cowpea: pre-flowering, flowering and post-flowering. Five sets of cowpea seeds were subjected to individual doses of gamma rays: 400 Gy, 450 Gy, 500 Gy, 600 Gy and 0 Gy. The seeds were sown to raise the M1 generation. The M1 generation seeds were collected and sown in the next season to raise the M2 generation. Leaf chlorophyll content was measured for M2 generation plants. Mean chlorophyll content for pre-flowering stage ranged between 38.9 ± 8.17 (control) and 64.2 ± 6.16 (400 Gy). Flowering stage mean chlorophyll content ranged from 48.3 ± 14.4 (600 Gy) to 99.4 ± 6.22 (450 Gy). Post-harvest chlorophyll mean content ranged between 13.1 ±0.98 (600 Gy) and 38.0 ±1.90 (400 Gy). There were significant differences in treatment effects for pre-flowering (P = 0.021), flowering (P = 0.005) and harvest (P = 0.000). At pre-flowering treatment, treatment 400 Gy scored a significant increase of 64 percent (P = 0.02) above the control. The optimum dose for useful induced mutation for increases in chlorophyll concentration in the cowpea was 400 Gy.

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

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.

Assessment of efficacy of mutagenesis of gamma-irradiation in plant height and days to maturity through expression analysis in rice.

Introduction of semi-dwarfism and early maturity in rice cultivars is important to achieve improved plant architecture, lodging resistance and high yield. Gamma rays induced mutations are routinely used to achieve these traits. We report the development of a semi-dwarf, early maturing and high-yielding mutant of rice cultivar 'Improved White Ponni', a popular cosmopolitan variety in south India preferred for its superior grain quality traits. Through gamma rays induced mutagenesis, several mutants were developed and subjected to selection up to six generations (M6) until the superior mutants were stabilized. In the M6 generation, significant reduction in days to flowering (up to 11.81% reduction) and plant height (up to 40% reduction) combined with an increase in single plant yield (up to 45.73% increase) was observed in the mutant population. The cooking quality traits viz., linear elongation ratio, breadthwise expansion ratio, gel consistency and gelatinization temperature of the mutants were similar to the parent variety Improved White Ponni. The genetic characterization with SSR markers showed variability between the semi-dwarf-early mutants and the Improved White Ponni. Gibberellin responsiveness study and quantitative real-time PCR showed a faulty gibberellin pathway and epistatic control between the genes such as OsKOL4 and OsBRD2 causing semi-dwarfism in a mutant. These mutants have potential as new rice varieties and can be used as new sources of semi-dwarfism and earliness for improving high grain quality rice varieties.

Rice strain selection for Mentik Wangi variety M3 generation resulting from 200 gray gamma irradiation

Mentik Wangi rice is a local variety that is quite popular with the people of Magelang. Mentik Wangi rice has a distinctive aroma and has a fluffier texture. The disadvantage of Mentik Wangi rice is the potential for lower production and the plants are too high. There is an effort done to overcome that problem were planted mutation through the utilization of gamma-ray irradiation. The aim of this research is to select M3 generation mutant rice resulting from 200 gray gamma-ray irradiation which has a short stems and has higher productivity and determine the performance of M3 generation mutant rice resulting from 200 gray gamma-ray irradiation. The observational data were analyzed descriptively and compared the performance of the Mentik Wangi M3 rice plants as a result of 200 gray gamma-ray irradiation with control plants through T-test analysis. The results showed that the performance of the M3 generation of Mentik Wangi rice was better than control plants, there were several strains with potential mutants traits based on the positive character of each individual. There are 24 Mentik Wangi M3 generation rice plants resulting from 200 gray gamma-ray irradiation plants which are selected based on short stem and have a high productivity.

Selection of M3 genotype short stem results of ray gamma 250 gray in Mentik Wangi rice

This study aims to determine the performance of Mentik Wangi rice at M3 generation mutant rice and select the M3 generation mutant plants as a result of 250 gray gamma irradiation which has short stems with high productivity. The research was carried out by planting 70 M2 lines of 250 gray gamma irradiation and control plants. The variables observed were plant height, total number of tillers, number of productive tillers, flowering age, harvest age, panicle length, number of seeds per panicle, panicle bush thickness index, weight of 100 seeds, weight of seeds per clump. Following observation, data were analyzed descriptively and statistically compared using t-test to determine the difference of M3 generation of fragrant Mentik rice from 250 gray gamma irradiation with control plants (without irradiation). There are 24 individual selected lines of M3 generation of fragrant Mentik Wangi mutants from 250 Gy gamma ray radiation which have a short stem character and have higher productivity than control plants.

Gamma Irradiation Enhanced Leaf Bioactive Components and Bioassay Parameters in M<sub>5</sub> Mulberry (<i>Morus</i> Sp.) Mutant

Gamma radiation is an effective tool for inducing genetic variation in plant characters. In the present experiment, M5 mulberry variety juvenile twigs were subjected to source Co60 gamma irradiation (1 kR - 10 kR) and mutants grown in triplicates in randomized block design to raise M1 and M2 generation. In M2 generation plants were subjected to phytochemical and bioassay tests. Silkworm rearing parameters and commercial characters of cocoons were recorded by feeding cross breed silkworms. Results show that M5 mutant leaves revealed significant variations in phytochemical constituents and moisture content. Bioassay tests recorded significant differences compared to control in M2 generation. Commercial characters like cocoon weight (1.41 g), shell weight (0.24 g), shell percentage (16.29 %), filament length (821.00 mts), renditta (8.2), denier (2.24) and effective rate of rearing (92.14 %) were increased. It is concluded that, gamma rays treatment enhances the mulberry plants leaf bioactive components, silkworm rearing and cocoon parameters and shows beneficial variants in M2 generation.

Impact of chemical mutagenesis using ethyl methane sulphonate on tepary bean seedling vigour and adult plant performance

Tepary bean is an important food legume. The genetic improvement of the crop is limited by a narrow genetic base but this genetic base can be broadened through induced mutagenesis. This study was designed to determine the (i) effects of chemical mutagenesis using ethyl methane sulfonate (EMS) on M1 seedlings (ii) the adult plant performance of M2 - M4 generations and (iii) the relationships between growth attributes at the seedling and adult plant stage. Mutagenized seed with varying doses of EMS was germinated at room temperature in order to raise M1 seedlings. There were highly significant (P < 0.01) differences due to dose effects among the seedlings. The highest LD50 (3.37 % EMS v/v) was observed for ‘Genotype 3’. Under field conditions, all the three factors influenced the plant performance. The results demonstrated the potential of EMS to induce genotypic variation and desirable agronomic traits in tepary bean.

Morphophysiological characteristics and yields of black rice (Oryza sativa L. Indica) induced by gamma rays 100 gy at m3 generation

Black rice is a popular local rice and has potential to be developed due to having benefit for health. The high crop posture, long time period for cultivating, and low productivity being limitation factors for black rice cultivation. Induced mutation using irradiation of gamma rays was expected to produce M3 mutant black rice that has better agronomic and physiological properties. The aims of this research were to examine morphophysiological characters by growth analysis and yield performance on M3 generation of local black rice varieties from Boyolali and Bantul as a result of 100 Gy gamma ray irradiation. The observational data were analysed descriptively and compared to local black rice varieties at a dose of 100 Gy with the control plants (without irradiation) through T-test analysis. The result showed morphophysiological character on M3 mutant black rice of Bantul varieties with dose 100 Gy based on growth analysis showed that the value of NAR, SLW, and CGR is high on 2–4 WAP. Bantul 0 Gy had a high increase of LAI on 2–4 WAP. Mutation that occurs on mutant M3 black rice at a dose of 100 Gy causing short plant height, but the yield components were still low.

Effect of gamma rays on chlorophyll content and ascorbic acid content of chilli in M1 generation

This study was performed by exposing the seeds of chilli (Capsicum annuum L.) cv. Phule Jyoti to gamma rays. The observations were made on chlorophyll content and Ascorbic acid content of chilli. The study revealed that ascorbic acid content decreased with an increase in dose/concentration of gamma rays as compared to control. Whereas in Chlorophyll content, maximum chlorophyll ‘a’ was observed in control, chlorophyll ‘b’ in lower dose of radiation and total chlorophyll in higher dose of gamma rays. Higher dose of radiation showed minimum chlorophyll ‘a’ and ‘b’ in M1 generation.

Screening for sheath blight resistance lines of rice (0ryza sativa L.) variety swarna by using of chemical mutagen

The present investigation was undertaken to identify the resistant lines amongst mutated rice population of the variety swarna induced by the sodium azide. The variability in disease reaction was observed among mutated rice lines. Out of the total 1000 mutant plants, 47 plants were screened that show low disease index, ranged between 0 to 5 were selected for next M generation. In M2 generation the selected genotypes show differences for disease index, only 12 lines shows resistance in M2 generation in field condition and in humidity chamber condition only 8 lines shows resistance. The number of resistance lines were decreases in humidity condition. Non significant variation in morphological expression and in yield attributes among the selected mutants was also reported. Thus, these lines could be considered a potential source for disease resistance against the sheath blight of rice and could be used further in the crossing programme for development of sheath blight resistant rice variety.

Mutation breeding in six rowed barley: Screening for stem rust resistance in certain induced mutants

During the induced mutagenesis in six rowed barley, 18 mutant lines were isolated from the M2 and M3 generations. Thirteen lines were induced by gamma rays, four by EMS and one by HA. These lines were screened for resistance to stem rust and loss assessment. Mutants were grown in field as well as in earthen pots. Artificial inoculums were provided in condition suitable for rust occurrence. Rust reaction was recorded according to modified Cabb’s scale and the rust intensity was classified as 0,5, 10, 25, 40 and 65 percent. Lines K50-38, K50-97, K50-102 and K40-73 were completely free from infection (0 %) and thought to be immune lines(I). K50-47, KE-4-47 and G50-47 exhibited traces of symptoms and placed under 5% intensity and were very resistant(R). Line KE2-63 exhibited 10% intensity and classified as moderately resistant(MR). Line K50-44; K40-11, K40-67, K30-91, KE3-75, G50-35 and GH3-32 showed 25% intensity of pustules and were put into moderately susceptible(MS) class. One line K30-82 showed 40% intensity and classified as susceptible(S), while line KE4-87 exihibited 65% intensity and was classified as heavily susceptible. Class 100 percent was not observed. Immune, very resistant and moderately resistant were identified as resistant lines and selected for further study, while moderately susceptible and heavily susceptible lines were identified as susceptible lines and discarded. Reduction in yield contributing traits i.e. number of grains/spike, test weight, seed yield and biological yield was also observed which was positively correlated with disease intensity.

Screening of M2 and M3 mutants of rice against bacterial leaf blight

A study was held at N. E. Borlaug Crop Research Centre during the kharif season of 2018-19, 2019-2020 and 2020-21. The seeds of various basmati and non-basmati lines were irradiated with ϒ rays at NBRI, Lucknow in 2018-19. The seeds were grown in the same year, the observed viability was less than 99.7 % (M1). The seeds were very meagre and so they were collected and grown in the kharif season of 2019-20. In this generation a screening test for examining BLB resistance was conducted. Resistant mutants were identified in the M2 generation. Similarly, the seeds obtained from M2 were grown as the M3 generation and again screening was done for the BLB. Some mutants were identified which revealed resistance and could be used in the future breeding program for the alleviation of disease susceptibility and catering resistance.

Development of EMS-induced Mutagenized Groundnut Population and Discovery of Point Mutations in the &lt;i&gt;ahFAD2&lt;/i&gt; and &lt;i&gt;Ara h 1&lt;/i&gt; 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.

Evaluation of Agro-morphological and Pollen Parameters of M&amp;lt;sub&amp;gt;2&amp;lt;/sub&amp;gt; Generation of &amp;lt;i&amp;gt;Capsicum annuum&amp;lt;/i&amp;gt; Exposed to Fast Neutron Irradiation (FNI)

Morphological and pollen parameter evaluation of M₂ generation of fast neutron irradiated capsicum annuum was carried out to assess the heritability of the useful traits observed in the M1 generation. Irradiated M₁ generation seeds at the Centre for Energy and Research Training (CERT), Ahmadu Bello University, Zaria using Fast Neutron Irradiation (FNI) from Americium-Beryllium source with flux 1.5x10⁴n.cm^-2s^-1 at 0, 30, 60, 90 and 120 minutes exposure periods were collected from the Department of Biological Sciences, Federal University of Technology Minna. The seeds were raise in the experimental garden of the Department in Randomised Complete Block Design (RCBD) and assess for morphological and pollen parameter. The results of the plant height indicated significant difference (P<0.05) across the treated plants at 2, 4, 6 and 8 weeks. The minimum height and number of leaf per plant at week 2 (3.53 cm and 6.25) and 4 weeks (11.80 cm and 13.85) was due to plant exposed to 30 minutes irradiated mutant. Maximum number of fruit and weight of fruit per plant was due to 60 minutes IEP with the value of 3.13 and 22.22 g respectively, while the minimum weight was due to 30 minutes exposure periods (13.89 g). Plants with 90 minutes exposure period recorded the highest percentage of pollen viability (96.78) and number of pollen production (79600.00).