Skip to main navigation menu Skip to main content Skip to site footer

Research Articles

Early Access

Ultrastructure of scutellum-induced callus deciphers stages of somatic embryogenesis and shoot regeneration in indica rice

DOI
https://doi.org/10.14719/pst.10708
Submitted
17 July 2025
Published
04-02-2026

Abstract

This study describes somatic embryogenesis from scutellar epithelial tissue-induced callus in indica rice cultivar PR126 followed by shoot regeneration. The highlighting feature was histological identification of distinct embryogenic regions revealing origin of embryogenesis from upper epidermal layer of callus cells. This layer comprised of clusters of small, compact and isodiametric pro-embryogenic cells with dense cytoplasm. These cells enlarged and differentiated, leading to the development of pro-embryos having distinctive protodermis that produced globular somatic embryos with an average frequency of 5.36 ± 2.89 % in 25 days old callus. The embryos formed discrete leaf primordium-like structures from the epidermal layer after 14 days of callus transfer on regeneration. The leaf primordia developed into shoot apices, followed by normal shoot formation at 77.06 ± 10.82 % frequency within 45 days. The study demonstrates that the embryogenic regions in calli are fewer; their early detection and promotion can lead to effective shoot regeneration. This approach can be used for trait improvement in recalcitrant commercial rice cultivars through genome editing.

References

  1. 1. Altpeter F, Springer NM, Bartley LE, Blechl AE, Brutnell TP, Citovsky V, et al. Advancing crop transformation in the era of genome editing. Plant Cell. 2016;28:1510–20. https://doi.org/10.1105/tpc.16.00196
  2. 2. Deo PC, Tyagi AP, Taylor M, Harding R, Becker D. Factors affecting somatic embryogenesis and transformation in modern plant breeding. South Pac J Nat Appl Sci. 2010;28(1):27–40. https://doi.org/10.1071/SP10002
  3. 3. Subban P, Kutsher Y, Evenor D, Belausov E, Zemach H, Faigenboim A, et al. Shoot regeneration is not a single cell event. Plants. 2020;10(1):58. https://doi.org/10.3390/plants10010058
  4. 4. Narciso JO, Hattori K. Genotypic differences in morphology and ultrastructures of callus derived from selected rice varieties. Philipp Sci Lett. 2010;3:59–65.
  5. 5. Abe T, Futsuhara Y. Efficient plant regeneration by somatic embryogenesis from root callus tissues of rice. J Plant Physiol. 1985;121(2):111–18. https://doi.org/10.1016/S0176-1617(85)80035-3
  6. 6. Vega R, Vásquez N, Espinoza AM, Gatica AM, Valdez-Melara M. Histology of somatic embryogenesis in rice (Oryza sativa cv. 5272). Rev Biol Trop. 2009;57:141–50. https://doi.org/10.15517/RBT.V57I0.21291
  7. 7. Mendoza AB, Futsuhara Y. Histological observations on plant regeneration in rice (Oryza sativa L.) calli. Jpn J Breed. 1992;42(1):33–41. https://doi.org/10.1270/jsbbs1951.42.33
  8. 8. Kruglova N, Zinatullina A, Yegorova N. Histological approach to the study of morphogenesis in callus cultures in vitro: a review. Int J Plant Biol. 2023;14(2):533–45. https://doi.org/10.3390/ijpb14020042
  9. 9. Ijaz B, Sudiro C, Hyder MZ, Malik SI, Farrukh S, Schiavo FL, et al. Histo-morphological analysis of rice callus cultures reveals differential regeneration response with varying media combinations. In vitro Cell Dev Biol Plant. 2019;55:569–80. https://doi.org/10.1007/s11627-019-09974-6
  10. 10. Murashige T, Skoog F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant. 1962;15(3):473–97. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
  11. 11. Sharma M. Genetic transformation of indica rice for sheath blight resistance using chitinase gene. PhD Thesis. Ludhiana: Punjab Agricultural University; 2021.
  12. 12. Pathania S, Lore JS, Kalia A, Kaur A, Sharma M, Mangat GS, et al. Conversion of sheath blight susceptible indica and japonica rice cultivars into moderately resistant through expression of antifungal β-1,3-glucanase transgene from Trichoderma spp. Transgenic Res. 2022;31(4):537–51. https://doi.org/10.1007/s11248-022-00318-6
  13. 13. Johansen DA. Plant microtechnique. New York: McGraw-Hill Book Co. Ltd.; 1940.
  14. 14. Ruzin SE. Plant microtechnique and microscopy. New York: Oxford University Press; 1999.
  15. 15. Verbelen JP, Kerstens S. Polarization confocal microscopy and Congo red fluorescence: a simple and rapid method to determine the mean cellulose fibril orientation in plants. J Microsc. 2000;198(2):101–07. https://doi.org/10.1046/j.1365-2818.2000.00691.x
  16. 16. Humphrey CD, Pittman FE. A simple methylene blue–azure II–basic fuchsin stain for epoxy-embedded tissue sections. Stain Technol. 1974;49(1):9–14. https://doi.org/10.3109/10520297409116929
  17. 17. Brisibe EA, Miyake H, Taniguchi T, Maeda E. Callus formation and scanning electron microscopy of plantlet regeneration in African rice (Oryza glaberrima Steud.). Plant Sci. 1992;83(2):217–24. https://doi.org/10.1016/0168-9452(92)90081-V
  18. 18. Falco MC, Mendes BMJ, Neto AT, Glória BA. Histological characterization of in vitro regeneration of Saccharum sp. R Bras Fisiol Veg. 1996;8(2):93–97.
  19. 19. Fransz PF, Schel J. Cytodifferentiation during the development of friable embryogenic callus of maize (Zea mays). Can J Bot. 1991;69(1):26–33. https://doi.org/10.1139/b91-005
  20. 20. Mostafz SB, Wagiran A. Efficient callus induction and regeneration in selected indica rice. Agronomy. 2018;8(5):77. https://doi.org/10.3390/agronomy8050077
  21. 21. Taylor MG, Vasil IK. The ultrastructure of somatic embryo development in pearl millet (Pennisetum glaucum; Poaceae). Am J Bot. 1996;83(1):28–44. https://doi.org/10.1002/j.1537-2197.1996.tb13871.x
  22. 22. Fransz PF, Schel JHN. Ultrastructural studies on callus development and somatic embryogenesis in Zea mays L. Berlin, Heidelberg: Springer; 1994.
  23. 23. Sangduen N, Klamsomboon P. Histological and scanning electron observations on embryogenic and non-embryogenic calli of aromatic Thai rice (Oryza sativa L. cv. Khao Daw Mali 105). Kasetsart J (Nat Sci). 2001;35:427–32.
  24. 24. Mangat BS, Pelekis MK, Cassells AC. Changes in the starch content during organogenesis in in vitro cultured Begonia rex stem explants. Physiol Plant. 2006;79(2):267–74. https://doi.org/10.1111/j.1399-3054.1990.tb06741.x
  25. 25. Appezzato-da-Glória B, Machado SR. Ultrastructural analysis of in vitro direct and indirect organogenesis. Rev Bras Bot. 2004;27:429–37.
  26. 26. Quiroz-Figueroa FR, Rojas-Herrera R, Galaz-Avalos RM, Loyola-Vargas VM. Embryo production through somatic embryogenesis can be used to study cell differentiation in plants. Plant Cell Tiss Org Cult. 2006;86(3):285–301. https://doi.org/10.1007/s11240-006-9139-6
  27. 27. Mantell SH, Matthews JA, McKee RA. Principles of biotechnology. Oxford: Blackwell Scientific Publications; 1985.
  28. 28. Razdan MK. Introduction to plant tissue culture. New Delhi: Oxford & IBH Publishing Co. Pvt. Ltd.; 2019.
  29. 29. Abe T, Futsuhara Y. Genotypic variability for callus formation and plant regeneration in rice (Oryza sativa L.). Theor Appl Genet. 1986;72(1):3–10. https://doi.org/10.1007/BF00261446
  30. 30. Islam MM, Haque ME, Alam S, Islam MA, Khalekuzzaman M, Sikdar B. Morphological and histological observation of embryogenic calli derived from immature embryo of BRRI dhan28 (Oryza sativa L.) variety. Res Plant Biol. 2013;3:21–27.
  31. 31. Nakano H, Maeda E. Shoot differentiation in callus of Oryza sativa L. Z Pflanzenphysiol. 1979;93(5):449–58. https://doi.org/10.1016/S0044-328X(79)80179-8
  32. 32. Indoliya Y, Tiwari P, Chauhan AS, Goel R, Shri M, Bag SK, et al. Decoding regulatory landscape of somatic embryogenesis reveals differential regulatory networks between japonica and indica rice subspecies. Sci Rep. 2016;6:1–15. https://doi.org/10.1038/srep23050
  33. 33. Kalhori N, Nulit R, Go R, Zulkifly S, Azizi P, Abiri R. Selection, characterization and somatic embryogenesis of Malaysian salt-tolerant rice (Oryza sativa cv. MR219) through callogenesis. Int J Agric Biol. 2017;19:157–63.
  34. 34. Yan LN, Xia LI, Dan WU. The comparison in tissue culture ability of mature embryo in different cultivars of rice. Agric Sci China. 2010;9:840–46. https://doi.org/10.1016/S1671-2927(09)60162-0
  35. 35. Khanday I, Santos-Medellín C, Sundaresan V. Rice embryogenic trigger BABY BOOM1 promotes somatic embryogenesis by upregulation of auxin biosynthesis genes. bioRxiv. 2020:2020.08.24.265025. https://doi.org/10.1101/2020.08.24.265025
  36. 36. Elhiti M, Stasolla C, Wang A. Molecular regulation of plant somatic embryogenesis. In vitro Cell Dev Biol Plant. 2013;49(6):631–42. https://doi.org/10.1007/s11627-013-9547-3
  37. 37. Sivanesan I, Nayeem S, Venkidasamy B, Kuppuraj SP, Rn C, Samynathan R. Genetic and epigenetic modes of the regulation of somatic embryogenesis: a review. Biol Futur. 2022;73:259–77. https://doi.org/10.1007/s42977-022-00126-3

Downloads

Download data is not yet available.