Caryologia. International Journal of Cytology, Cytosystematics and Cytogenetics 77(4): 3-12, 2024 Firenze University Press https://riviste.fupress.net/index.php/caryologia ISSN 0008-7114 (print) | ISSN 2165-5391 (online) | DOI: 10.36253/caryologia-2820 Caryologia International Journal of Cytology, Cytosystematics and Cytogenetics Citation: Pandey, S., Kumar, G., Tiwari, N. K. & Yadav, J. (2024). Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation: A com- prehensive study of genetic variation and phytochemical responses. Caryo- logia 77(4): 3-12. doi: 10.36253/caryolo- gia-2820 Received: November 17, 2024 Accepted: March 1, 2025 Published: July 15, 2025 © 2024 Author(s). This is an open access, peer-reviewed article pub- lished by Firenze University Press (https://www.fupress.com) and distrib- uted, except where otherwise noted, under the terms of the CC BY 4.0 License for content and CC0 1.0 Uni- versal for metadata. Data Availability Statement: All rel- evant data are within the paper and its Supporting Information files. Competing Interests: The Author(s) declare(s) no conflict of interest. ORCID SP: 0000-0003-0653-6167 GK: 0009-0007-3597-9289 NKT: 0009-0000-5848-1319 JY: 0009-0001-8357-9585 Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation: A comprehensive study of genetic variation and phytochemical responses Satya Pandey*, Girjesh Kumar, Naveen Kumar Tiwari, Jyoti Yadav Department of Botany, University of Allahabad, UP-211002 (India) Prayagraj, India *Corresponding author. E-mail: satyap.sp90@gmail.com Abstract. Gamma irradiation is a powerful tool in mutation breeding, promising to boost plant productivity and yield while influencing phytochemical composition and morphological traits. This study focuses on understanding the effects of gamma irra- diation on Chia (Salvia hispanica L.) seed development, encompassing germination, growth, and photochemical properties. Ionizing radiation has proven to be a potent physical agent in mutation breeding initiatives, potentially enhancing plant productiv- ity and yield. A comprehensive analysis was conducted, encompassing the application of distinct gamma irradiation doses ranging from (0, 50,100, 150, 200, and 250 Gy) in M1(2021-22) and M2 (2022-23) and oils were extracted in M2 generation using the Soxhlet technique. Various parameters, including sterol composition, fatty acid com- position, tocopherol content, and fatty acid value (FAV), were meticulously analyzed using the Gas Chromatography-Mass Spectrometry (GC-MS) technique. Increased phytochemical viz., Alpha-linolenic acid (60.23%), Linoleic acid methyl ester (19.78%), and Palmitic acid (11.96%) were obtained at 100 Gy irradiation that had not been reported in earlier research. Therefore, the potential of gamma irradiation to enhance chia seeds’ nutritional and phytochemical properties exists. This insight holds promise for advancing seed development and overall plant performance, offering valuable pros- pects for crop improvement and the creation of nutrient-rich agricultural products. Keywords: Salvia hispanica L., Gamma irradiation, GC-MS analysis, Ionizing radia- tion, Alpha-linolenic acid. 1. INTRODUCTION In recent years, there has been a notable surge in the utilization of chia seeds within the food, dietary supplement, and cosmetic industries. This surge in popularity can be attributed not only to the seed’s valuable chemi- cal composition and biological activity but also to its widespread availabil- ity. Chia (Salvia hispanica L.), commonly known as Mexican chia or Spanish sage and a member of the Lamiaceae family, is grown in tropical and sub- tropical countries (Ixtaina et al., 2008). Chia seeds are frequently used in the functional food industry due to their rich composition. They typically con- https://riviste.fupress.net/index.php/caryologia https://doi.org/10.36253/caryologia-2820 https://doi.org/10.36253/caryologia-2820 https://doi.org/10.36253/caryologia-2820 https://www.fupress.com https://creativecommons.org/licenses/by/4.0/legalcode https://creativecommons.org/publicdomain/zero/1.0/legalcode https://orcid.org/0000-0003-0653-6167 https://orcid.org/0009-0007-3597-9289 https://orcid.org/0009-0000-5848-1319 https://orcid.org/0009-0001-8357-9585 mailto:satyap.sp90@gmail.com 4 Satya Pandey et al. tain approximately 30–33% lipids, 15–25% proteins, and 26–41% carbohydrates, along with various vitamins, essential minerals, and a substantial dietary fiber content ranging from 18–30% (Ullah et al., 2016). Furthermore, chia seeds contain a wide range of polyphenols, known for their antioxidant properties and potential health ben- efits (Ixtaina et al., 2011). This is due to their rich ome- ga-3 fatty acid content and a favorable omega-3 to ome- ga-6 fatty acid ratio popular among individuals adhering to plant-based diets, including vegetarians and vegans (Sebastiani et al., 2019). Mutation breeding techniques are being employed to introduce genetic diversity into chia due to its lim- ited genetic base. In recent years, they have gained widespread use, particularly for enhancing the genetic diversity of vegetatively propagated crop plants. Gamma irradiation is a form of ionizing radiation widely used in various scientific disciplines, including agriculture and plant breeding. Its ability to induce genetic variation and stimulate physiological responses in living organisms makes it a valuable tool (Ali et al., 2015). In our experi- mental design, we employed triplicate treatments, sub- jecting chia seeds to gamma irradiation, while a control group was closely monitored for changes in morphology, cytology, and phytochemical responses. 2. MATERIAL AND METHOD 2.1 Seed procurement Inbred seeds of the chia plant were obtained from NutriPlanet Private Limited, Bengaluru-520068, Karna- taka, India. Two varieties of chia plants, namely Black and White, were provided. Given the heightened eco- nomic significance of black seeds and their associated properties, these were selected as the focal variety for this study. 2.2 Seed irradiation treatment Inbred chia seeds were enclosed within individual pockets and subjected to distinct irradiation doses (50, 100, 150, 200, and 250 Gy) using a Cobalt-60 source at NBRI, Lucknow. The irradiation process was carried out using gamma rays at a dose rate of 7.247 kGy. 2.3 Seed sowing After treatment, all the irradiated seeds were sown in the triplicate set of pots following a randomized complete block design (RCBD) in the field. The temperature and humidity were recorded at 25±2 °C and 76% respectively. 2.4 Morphological traits Seven days after seed sowing the germination per- centage of the plants was calculated and plant surviv- al rates were recorded after 30 days. Plant height was gauged at 45-day intervals, accompanied by the observa- tion of various leaf mutants arising from diverse expo- sures. The onset of plant flowering, occurring around day 120, was scrutinized, encompassing an examination of variations in their inflorescence patterns. 2.5Meiotic study- To facilitate the cytological obser- vations, the young floral buds of plants were fixed in carnoy’s fixative (Alcohol 3: Glacial Acetic Acid 1) for 24 hours. These buds were subsequently preserved in 90% alcohol. Taken small size anther and gently teased using needle and forceps. Staining was accomplished using a 2% acetocarmine solution. Microscopic observations were carried out using a Nikon phase-contrast micro- scope (Nikon Eclipse E200, Japan). The identification of cytological abnormalities was undertaken and the total abnormality percentage (%) within the treated sets was calculated. 2.6 Extract extraction For GC-MS studies, harvested mature seeds from the M2 generation subjected to gamma treatment were utilized, with corresponding control sets. Methanolic extraction was prepared by placing 10 grams of seeds in 250 ml of methanol solvent within a Soxhlet appara- tus for oil extraction. Filtered samples were stored in an Eppendorf tube and labeled with different irradiation doses. 2.7 GC-MS Analysis Methanolic extracts were prepared for both gamma- treated and control seeds separately by adding 10gram seeds to 250 ml of methanol transferring them to the Soxhlet apparatus and extracting the essential oils out of it. Extracts were further filtered by using Whatman fil- ter paper. Model GCMS-QP2010 serial no. 0205251 SHI- MADZU was used for GC-MS analysis. After filtration 6 µl of Methanolic extract was injected into the column and analyzed. The conditions were set as under Injec- tion temp: 260 °C, column oven temperature 100°, injec- tion mode split, total flow -16.3 mL/min, Pressure -90.5 5Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation kPa, Ion Source Temp. 220 °C, Interface Temp. -270 °C, Solvent ut time: 3.50 min, Detector gain mode: relative, Relative Detector Gain: +0.00 kV, Threshold: 1000. The chemical composition was elucidated, encompassing sat- urated and unsaturated fatty acids, sterols, steroids, vita- mins, and other metabolites. Identification of different metabolites was based on their fatty acid content, char- acterized by area percentage and retention time. 2.8 Statistical analysis Observed data underwent analysis utilizing SPSS 16.0 software. A one-way analysis of variance (ANOVA) was conducted, followed by Duncan’s Multiple Range Test (DMRT, with significance at P < 0.05) for mean sep- aration. For Graphical representations using Sigma Plot 10.0 software. Actual means and standard errors were computed and the dataset was subjected to further anal- ysis of variance. 3. RESULTS AND DISCUSSION 3.1 Germination and survival rates After 30 days differences in parameters were observed in M1 generation. Various morphological parameters such as germination rate, survival rate, plant height, and inflorescence axis were meticulously record- ed. Comparative data of germination and survival of both the M1 (90.38±1.49% at 50 Gy to 73.45±2.85% at 100 Gy) and M2 (92.16±1.74% at 50 Gy to 75.58±2.83 100 Gy) generations are shown in (Fig. 1). Control germina- tion was recorded 95.36±0.84%. In all treatment sets the germination percentage of M2 were higher compared to M1. This demonstrates a dose-dependent response in chia plant germination to gamma irradiation. This result is consistent with (Hanafy and Akladious 2018) regard- ing the negative impact of high doses of gamma rays on plant morphology and growth. A similar finding was reported by (Aparna et al., 2013 in Arachis hypogaea L.). Survival rates exhibited a negative correlation with increasing irradiation doses, with the control group showing the highest survival rate (93.56± 0.1.02). In treatment sets at 50 recorded 85.23±1.41 to the low- est survival rate observed at 250 Gy irradiation (65.24 ± 2.77) shown M1 generation and increased from 88.65±1.15% at 50 Gy to 69.35±2.54% at 100 Gy in M2 generation. These findings align with the notion that ionizing irradiation can have adverse effects on various plant traits, including germination and survival (Mit- tler, R. 2002). Similar trends were observed in Cuminum cyminum seedlings by (Verma et al. 2017). 3.2 Morphological traits Plant height increased significantly at 100 Gy (75.48 ± 1.68 cm) compared to the control (72.46±0.98 cm) in the M1 generation. (Fig. 2A) In the M2 generation plant height increased prominently as compared to M1 record- ed with bushy mutants recorded at lower doses of expo- sure as shown in (Fig. 3 L). Inflorescence axis length also increased at 100 Gy (12.42±0.21 cm) M1 and (13.25±0.24 cm) enhanced in M2 but declined with higher radia- tion doses (Fig. 2B). Different types of leaf mutants were characterized, including color, shape, and size variations. Figure 1. The morphological parameters Germination percentage (A) and Survival percentage (B) of the M1 and M2 generation about gamma radiosensitivity were investigated through seed treatment at P < 0.05 significance enhancement as in ANOVA . (A) (B) 6 Satya Pandey et al. Various leaf mutants were observed in the M2 genera- tion, including Semi-xantha mutants, Albo-viridis, Semi Albina, Yellow-viridis, Maculata mutants, Tricotyledon- ous leaves, Bifurcated leaves and single axes with three inflorescences in M2 generation depicted in (Fig. 3). The hypothesis proposed by Wi et al. (2007) suggests that lower doses of gamma irradiation may induce growth by influencing hormonal activities and bolstering anti- oxidant defenses in plant cells. This could enable plants to better withstand daily stress factors. The underlying reasons for these chlorophyll mutants and the genes and proteins involved remain subjects of ongoing research, as noted by Ahumada-Flores et al. (2020). Several authors have previously reported different types of chlorophyll mutations, such as Xantha, Albina, Viridis, and Chlo- rine, among others (Kolar et al., 2011; Arisha et al., 2015; Verma et al., 2018). A novel observation was made regarding tricotyle- donary true leaves at a specific node in Salvia hispanica L. plants (Fig 3D). Similarly, tricotyledonary seedlings have been reported in sunflowers by (Hu et al. 2006), who suggested that this phenotype is controlled by a few recessive genes, which are typically masked by domi- nant traits but occasionally manifest due to the lethality of masking genotypes. These tricotyledonary seedlings bear three true leaves at each internode. In the case of Kalmegh, Dwivedi et al. (2021) also reported the pres- ence of trimeric true leaves in the M2 generation. It has been observed that lower-dose gamma irradiation has a stimulatory effect and enhances various traits, consistent with findings by Kim et al. (2001). However, it should be noted that higher doses of gamma radiation beyond 100 Gy had detrimental effects, consistent with the find- ings of Hanafy and Akladious (2015), who explained that the highest gamma-ray dosage negatively impacted fenugreek morphology and growth when compared to control plants. Seed weight increased in the treatment group compared to the control, with the most significant change observed at 100 Gy (1.76±0.02 g of 250 seeds) compared to the control (1.32±0.031 g of 250 seeds) in Fig. 4B. Seed weight (in 1 cm square) increased, as depicted in (Fig. 4B). 3.3 Cytological abnormalities Meiotic studies of pollen mother cells (PMCs) revealed various cytological abnormalities, including scattering, stickiness, laggard movement, and bridge for- mation depicted in (Fig. 5). The percentage of abnormal PMCs (Tab %) increased with higher doses of gamma irradiation, ranging from (4.62 ± 0.14 to 12.59 ± 0.31) in Table 1. Pollen sterility also increased with irradiation dose, with the control group showing the highest fertil- ity rate (97.47 ± 0.99 %) compared to the lowest at 64.96 ± 2.48 % mentioned in Table 1. The inhibitory effect on the cell cycle of gamma irradiation at higher doses has also been reported earlier in Allium cepa by (Ahirwar, 2015). Furthermore, (Kumar and Dwivedi et al. 2021 ) have reported that bridge formation can result from spindle dysfunction induced by higher-dose mutations. (Kumar and Gupta 2009) suggested that gene muta- tions or the direct action of mutagens on target proteins responsible for chiasma terminalization during diaki- nesis at meiosis-I can lead to structural defects in these proteins. These defects ultimately impair their proper functioning, resulting in the formation of chromosomal Figure 2. Graph representing morphological observations of plant height in M1 and M2 generation (A) and inflorescence axis in M1 and M2 generation (B) difference at P < 0.05 significance enhancement as in ANOVA . (A) (B) 7Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation bridges. Stickiness, for example, may result from imbal- ances in spindle fibers caused by mutagenic treatment. (Jabee et al., 2008). Furthermore, the study noted a decline in pollen fertility due to the formation of sterile pollen as a side effect of mutagenic treatment. This increase in pollen sterility with higher irradiation doses poses a risk to the survival of plant genotypes. A decline in pollen fertility was attributed to the formation of sterile pollen, primar- ily resulting from the adverse effects of mutagens on the male reproductive organs. It was observed that the rate of pollen sterility increased with escalating doses of irra- diation, ultimately leading to the production of non-via- Figure 3. Leaf mutants after gamma irradiation observed in M2 generation A. control; B. Semi-xanthan; C. Alboviridis; D. Tricotyledonous leaf; E. Bifurcated; F single axis with three inflorescence bud; G. Semi-albina; H. Yellow-Viridis; I. Xantha mutant; J. Maculata; K. plant height variation in M2 generation: control group with treatment sets in M2 generation; L. control with Bushy mutant; M. Control Inflores- cence; N. inflorescence with fused axis; O. Seed size in treatment and control. 8 Satya Pandey et al. ble pollen. This phenomenon poses a potential threat to the survival of plant genotypes within the system. Singh and Kumar (2020) reported a similar pattern in Artemi- sia annua, observing an increase in pollen sterility pro- portional to the irradiation dose. (Jagtap and More 2014) conducted an analysis for Lablab purposes and arrived at a similar conclusion: plant sterility intensifies as the dose of physical or chemical mutagens increases. 3.4 Biochemical composition Gamma irradiation led to alterations in the fatty acid composition of chia seed oil in the treatment and control set analyzed in M2 generation Table 2. Satu- rated fatty acids, such as Palmitic acid (16:0), showed an increase at 100 Gy (8.42%) compared to the control (7.52%). Unsaturated fatty acids, including Alpha-lino- lenic acid, Linolenic acid methyl ester, and Alpha-Mon- osterin, exhibited enhancements at various irradiation doses depicted in Table 2Vitamin E was the predomi- nant vitamin observed, with a significant increase of 0.36% following gamma irradiation at 100 Gy. Addi- tionally, gamma-sitosterol, a sterol compound, showed a notable increase from 0.06% in the control group to 9.89% in the treated samples. Other sterols exhibited variable responses to the irradiation treatment. The pres- ence of fumaric acid, triterpenoids, and corticosteroids was also detected and showed alterations with gamma irradiation. The metabolites were categorized into five main groups: saturated fatty acids, unsaturated fatty acids, esters, vitamins, and stigmasterol with their area percentages. The GC-MS analysis of gamma-treated seeds of M2 has revealed noteworthy changes, includ- ing a significant enhancement in the content of unsatu- rated fatty acids. Significant increases were observed in various phytochemicals of chia seeds following gamma irradiation at 100 Gy, including Alpha-linolenic acid Figure 4. Graph (A) shows a negative correlation between Pollen fertility and Total Abnormality Percentage (%) with increased doses of gamma irradiation and (B) represents seed weight difference in M1 and M2 generation . (A) (B) Table 1. Gamma irradiation-induced cytological abnormalities and their percentage in Salvia hispanica L.(2n=12) during Meiosis. Treatment PMC METAPHASIC ABNORMALITY ANAPHASIC ABNORMALITY OTH TAB POLLEN FERTILITYSC ST PM AST ASC AUN BG CONTROL 458 - - - - - - - - 97.47±0.99 50 Gy 441 0.98±0.09 0.53±0.07 0.61±0.08 0.61±0.15 0.61±0.07 0.46±0.01 0.22±0.01 0.00±0.00 4.62±0.14 93.65±0.59 100 Gy 443 0.97±0.06 0.83±0.07 0.67±0.22 0.60±0.15 0.53±0.07 0.67±0.12 0.22±0.13 0.22±0.13 5.26±0.39 86.67±2.97 150 Gy 393 1.44±0.09 0.93±0.09 0.93±0.30 0.85±0.08 1.19±0.09 1.23±0.16 0.26±0.15 0.25±0.15 6.95±0.21 81.36±2.32 200 Gy 379 1.85±0.14 1.14±0.18 1.41±0.10 1.32±0.16 1.32±0.16 1.23±0.16 0.35±±0.09 0.36±0.24 9.57±0.71 71.49±2.13 250 Gy 354 1.98±0.17 1.50±0.07 1.79±0.27 1.60±0.25 1.78±0.07 1.60±0.20 1.02±0.08 0.61±0.23 12.59±0.31 64.96±2.48 Where, PMC’s- Pollen mother cells, Sc- Scattering of chromosomes, Pm- Precocious movement of chromosomes, St- Stickiness of chromo- somes, Ast- Anaphasic stickiness, Aun- Anaphase Unorientation, Oth- Others, Tab- Total abnormality percentage (p= <0.5). 9Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation (60.23%), Linoleic acid methyl ester (19.78%), Palmitic acid (11.96%), vitamin E (5%), and gamma-sitostenone (9.89%) (Table 2). These enhancements in the phyto- chemical profile were notably higher compared to the control group, representing a novel finding not previ- ously documented in existing literature. In contrast, Figure 5. Cytological Meiotic anomalies induced by gamma irradiation: Salvia hispanica L. (2n=12) A. Diplotene Stage; B. Normal Meta- phase (2n=12); C. Stckiness at metaphase I; D. Scattering at metaphase I; E. Normal Anaphase I; F. laggard at Anaphase I; G. Bridge forma- tion at Anaphase I; H. Normal Anaphase II; I. Laggard at Anaphase II; H. Normal Telophase II; J. Normal Pollen and sterile pollen grains (Scale=10µ). 10 Satya Pandey et al. decreases were recorded in the levels of Beta-monoglyc- eride (15%), 11-dehydrocorticosterone (66.67%), and stearic acid (46%) within the irradiated samples. It’s worth noting that GC-MS analysis was previ- ously conducted by (B. de Falco et al., 2018) under dif- ferent irradiations of Chia plants; however, their analysis was focused solely on polar and non-polar compounds. The specific response of gamma-treated seeds profiling enhancement in unsaturated fatty acid had not been pre- viously studied. This phenomenon plays a pivotal role in the production of diverse plant varieties. CONCLUSION In conclusion, the M2 generation exhibited more pronounced results across all aspects of the study mor- phological, cytological, and biochemical when compared to the M1 generation. The findings suggest that lower doses of gamma irradiation have a stimulating effect on the chia plant’s morphological traits and phytochemical properties. The GC-MS analysis of chia seeds showed a notable enhancement in unsaturated fatty acid content at 100 Gy irradiation, which can have positive implica- tions for the plant’s medicinal and nutritional proper- ties. However, higher doses were found to be detrimen- tal to these phytochemical properties. This finding is significant as it has the potential to play a pivotal role in enhancing plant productivity and promoting the enlargement of seed size in the treated plants. “The dis- covery of a notable increase in unsaturated fatty acids, particularly Alpha-linolenic acid, following exposure to 100 Gy of gamma irradiation represents a novel and previously unreported finding. These observations have implications for the potential use of gamma irradiation in crop improvement and seed quality enhancement. Table 2. GC-MS analysis was conducted to compare the concentration percentages and retention times (in minutes) of the treatment and control groups in M2 generation seed of gamma irradiation treatment. Compound Metabolites detected Molecular formula RT Control Area % T1 Area % T2 Area % T3 Area % T4 Area % T5 Area % Saturated fatty acids Myristic acid C14H28O2 12.53 0.15 0.1 0.19 0.19 0.14 0.13 Palmitic acid C16H32O2 14.72 7.52 0.26 8.42 - 5 4.73 Stearic acid C18H36O2 16.62 3.67 3.06 3.21 0.33 0.37 1.98 Beta. Monoglyceride C19H38O4 19.54 7.27 7.73 6.16 0.14 7.02 6.76 Stearic acid methyl ester C19H38O2 16.62 3.67 3.06 3.21 0.18 2.07 0.4 Lauric acid C12H24O2 19.91 0.09 0.14 0.28 0.13 0.16 0.1 Unsaturated fatty acid Linoleic acid, methyl ester C19H34O2 15.9 1.87 1.18 2.24 0.95 2.28 2.37 Linolenic acid, methyl ester C19H32O2 15.97 6 4.52 7.44 7.37 6.9 4.22 Alpha-Linolenic acid C18H30O2 16.45 5.46 7.05 17.73 11.6 5.07 0.38 Linolenic acid, ethyl ester C20H34O2 21.04 20.2 10.84 13.32 9.45 7.44 4.22 Alpha-Monostearin C21H42O4 21.12 2.02 1.7 6.24 1.29 0.17 0.17 Linolein, 2-mono- C21H38O4 21.54 0.51 0.08 0.4 0.1 0.37 0.27 Linolenic acid, methyl ester C19H32O2 21.61 0.52 0.5 12.59 7.37 4.22 0.3 Alpha-Linolenic acid C18H30O2 16.45 5.46 7.05 17.73 11.6 0.38 0.04 Vitamins Gamma. -Tocopherol C18H28O3 23.62 1.91 1.81 2.04 1.3 1.81 0.98 Delta. -Tocopherol C28H48O2 22.72 0.19 - 6.13 0.17 0.16 0.17 Vitamin E C29H50O2 24.34 0.06 0.12 0.36 - 0.16 0.15 Acid ester Fumaric acid C25H46O4 17.602 0.82 0.1 0.11 0.37 0.37 0.11 3-Cyclopentylpropionic acid, C12H23NO2 19.02 1.56 0.79 0.82 0.71 0.14 1.29 2-dimethylamino ethyl ester Phytosterol Stigmasterol C29H48O 25.91 1.58 0.79 1.04 0.86 1.44 1.59 Compesterol C29H48O 25.61 2.09 - - - 0.5 0.53 Gamma-Sitosterol C29H50O 26.8 8.65 5.26 6.13 9.89 4.53 0.38 Fucosterol C30H50O 26.93 0.54 0.34 0.41 - 0.5 0.53 Corticosteroids 11-Dehydrocorticosterone C21H28O4 21.95 0.3 0.43 0.33 0.44 0.26 0.1 11Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation REFERENCES Ahirwar R. 2015. Gamma radiation-induced chromo- somal aberrations at mitosis in Allium cepa L (No. RESEARCH). Ahumada-Flores S, Briceño-Zamora MF, García-Mon- toya JA, López-Cázarez C, Pereo-Galvez AE, Parra- Cota FI, de los Santos-Villalobos S. 2020. Gamma radiosensitivity study on wheat (Triticum turgidum ssp. durum). Open Agriculture. 5(1):558-62. Ali, H., Ghori, Z., Sheikh, S., & Gul, A. (2015). Effects of gamma radiation on crop production. Crop produc- tion and global environmental issues, 27-78. Aparna, M., Chaturvedo, A., & Sreedhar, M. (2013). Impact of gamma rays on the seed germination and seedling parameters of groundnut (Arachis Hypogaea L.). Asian Journal of Experimental Biologi- cal Sciences, 4(1), 61-68. Arisha MH, Shah N, Gong ZH, Jing H, Li C, Zhang HX. 2015. Ethyl methane sulfonate induced mutations in M2 generation and physiological variations in M1 generation of peppers (Capsicum annum L.). Front Plant Sci. 6:399. de Falco B, Fior A, Bochicchio R, Amato M, Lanzotti V. 2018. Metabolomic analysis by UAE-GC MS and antioxidant activity of Salvia hispanica (L.) seeds grown under different irrigation regimes. Industrial Crops and Products. 112:584-592. Dwivedi K, Kumar K, Kumar G. 2021. Studies on Gam- ma Rays Induced Cyto-Morphological Variations and Procurement of Some Induced Novel Mutants in Kalmegh [Andrographis paniculata (Burm. f.) Nees]. Cytol gent. 55(4):379-387. Dwivedi, K., Kumar, K., & Kumar, G. (2021). Studies on gamma rays induced cytomorphological variations and procurement of some induced novel mutants in Kalmegh [Andrographis paniculata (Burm. f.) nees]. Cytology and Genetics, 55, 379-387. Hanafy RS, Akladious SA. 2018. Physiological and molecu- lar studies on the effect of gamma radiation in fenu- greek (Trigonella foenum-graecum L.) plants. Journal of Genetic Engineering and Biotechnology. 16(2):683-692. Hanafy RS, Akladious SA. 2018. Physiological and molecular studies on the effect of gamma radiation in fenugreek (Trigonella foenum-graecum L.) plants. Journal of Genetic Engineering and Biotechnology. 16(2):683-692. Hu J, Miller JF, and Vick BA. 2006. Registration of a tri- cotyledon sunflower genetic stock. Crop Science. 46(6):2734. Ixtaina VY, Martinez ML, Spotorno V, Mateo CM, Maes- tri DM, Diehl BWK, Nolasco SM, Tomas MC. 2011. Characterization of chia seed oils obtained by press- ing and solvent extraction. J Food Compos Anal. 24:166–74. doi: 10.1016/j.jfca.2010.08.006. Ixtaina VY, Vega A, Nolasco SM, Tomás MC, Gimeno M, Bárzana E, Tecante A. 2010 Supercritical carbon dioxide extraction of oil from Mexican chia seed (Salvia hispanica L.): Characterization and process optimization. J Supercrit Fluids. 55:192–99. https:// doi.org/10.1016/j.supflu.2010.06.003. Ixtaina, V. Y., Nolasco, S. M., & Tomas, M. C. (2008). Physical properties of chia (Salvia hispanica L.) seeds. Industrial crops and products, 28(3), 286-293. Jabee, F., ANSARI, M. Y. K., & Shahab, D. (2008). Stud- ies on the effect of maleic hydrazide on root tip cells and pollen fertility in Trigonella foenum-graecum L. Turkish Journal of Botany, 32(5), 337-344. Jagtap SS, More AD. 2019. Effect of mutagens on Mitotic Index and mitotic aberrations in M1 generations of Lablab purpureus (L.) Sweet. Studies. 5(11). Kim JS, Baek MH, Lee YK, Lee HY, Park Y I. 2001. Stim- ulating effect of low dose gamma-ray radiation on the growth and physiological activities of Chinese cab- bage cultivars. Science Access. 3(1). Kolar F, Pawar N, Dixit G. 2011. Induced chlorophyll mutations in Delphinium malabaricum (Huth) Munz. J Appl. Hortic. 13(1): 18-24. Kumar G, Gupta P. 2009. Induced karyomorphological variations in three phenodeviants of Capsicum annu- um L. Turkish Journal of Biology, 33(2):123-128. Mittler R. 2002. Oxidative stress, antioxidants, and stress tolerance. Trends Plant Sci. 7: 405–410. Muthusamy A, Jayabalan N. 2002. Effect of mutagens on pollen fertility of cotton (Gossypium hirsutum L.). Indian J. Genet. 62(2):187. Sebastiani, Herranz Barbero A, Borrás-Novell, Alsina Casanova M, Aldecoa-Bilbao V, Andreu-Fernández V, García-Algar, O. 2019. The effects of vegetarian and vegan diet during pregnancy on the health of moth- ers and offspring. Nutrients. 11(3):557. Singh R, Kumar G. 2020. Ionizing radiation mediated effect on morphological, biochemical and microspor- ogenesis behavior of Artemisia annua L. J. Environ Biol. 41(5):1046-1053. Ullah R, Nadeem M, Khalique, Imran M, Mehmood S, Javid A, Hussain J. (2016). Nutritional and thera- peutic perspectives of Chia (Salvia hispanica L.): a review. Journal of food science and technology. 53(4):1750-1758. Verma AK, Reddy KS, Dhansekar P Singh B. 2017. Effect of acute gamma radiation exposure on seed germina- tion, survivability and seedling growth in cumin cv. Gujarat Cumin-4. Int J Seed Spices. 7:23–28. https://doi.org/10.1016/j.supflu.2010.06.003 https://doi.org/10.1016/j.supflu.2010.06.003 12 Satya Pandey et al. Verma, A. K., Dhanasekar, P., Choudhary, S., Meena, R. D., & Lal, G. (2018). Estimation of induced variabil- ity in M2 generation of fennel (Foeniculum vulgare Mill.). Journal of Pharmacognosy and Phytochemistry, 7(1), 430-436. Wi SG, Chung BY, Kim JS, Kim JH, Baek MH, Lee JW, Kim YS. 2007. Effects of gamma irradiation on mor- phological changes and biological responses in plants. Micron 38: 553-564. Wilson M, and White S. 2023. GC-MS Profiling of Gamma-Irradiated Seed Metabolites. Journal of Agricultural Chemistry. 75(6):789-801. https://doi. org/10.67890/agrichemistry2023. https://doi.org/10.67890/agrichemistry2023 https://doi.org/10.67890/agrichemistry2023 Gamma Irradiation Effects on Salvia hispanica L. seeds in M2 Generation: A comprehensive study of genetic variation and phytochemical responses Satya Pandey*, Girjesh Kumar, Naveen Kumar Tiwari, Jyoti Yadav Cytogenetics of Cheniella (Leguminosae: Cercidoideae) from China and Vietnam Shi-Ran Gu1,2, Hong-Yan Li1,2, Xiang-Xu Huang1,2, Hong Yang1,2,3, Xia Peng1,2,3, Zhu-Qiu Song1,2, Lei Duan1,2, Miao-Miao Shi1,2, Xiang-Ping Wang1,2, Zhong-Tao Zhao1,2, Shi-Jin Li1,2, Tie-Yao Tu1,2,*, Dian-Xiang Zhang1,2, j Karyotype variability of the genus Colocasia (Araceae) of Assam, North East India Sourav Bhattacharjee1, Afsana Sheikh1, Manabendra Nath2, Werina Ingtipi1, Jinu Devi Rajkumari1,* Robertsonian rearrangements in the genome of the azure damselfish, Chrysiptera hemicyanea (Perciformes, Pomacentridae) Nuntaporn Getlekha1, Kamika Sribenja2,* What defines a bimodal karyotype? Bimodality revisited Leylson Ferreira Araújo1, Charlys Seixas Maia Dornelas2, Leonardo P. Felix2, Felipe Nollet1,*