In Vitro Cellular & Developmental Biology - Plant

, Volume 55, Issue 6, pp 668–677 | Cite as

Genetic homogeneity and high shoot proliferation in banana (Musa acuminata Colla) by altering medium thiamine level and sugar type

  • Marwa Talaat El-Mahdy
  • Muhammad YoussefEmail author


To enhance the multiplication rate in Musa acuminata Colla (banana; ‘Grand Nain’) organogenesis, higher amounts of thiamine along with different sugar types and concentrations were evaluated at the proliferation phase. Thiamine at 1, 10, 50, 100, and 200 mg L−1 was compared with 0.1 mg L−1 thiamine found in conventional Murashige and Skoog (MS) medium. Maximum proliferation of banana was induced with 100 mg L−1 thiamine. Additionally, 15, 30, and 45 g L−1 sucrose, glucose, fructose, and sorbitol combined with regular and optimal levels of thiamine were tested. Glucose at 30 g L−1 most improved shoot proliferation alone and enhanced shoot proliferation further, when combined with 100 mg L−1 thiamine, followed by sucrose and fructose, whereas sorbitol completely inhibited growth and caused tissue browning. All evaluated vegetative traits were significantly affected by sugar type and concentration, and thiamine levels, unlike the photosynthetic pigments. Moreover, genetic stability of the plants recovered from the enhanced protocol was confirmed by inter-simple sequence repeats (ISSR) and randomly amplified polymorphic DNA (RAPD) analysis. A total of 230 bands generated by both marker types were monomorphic for the randomly selected regenerated plants, compared with their mother plant. Thus, the proliferation medium supplemented with 30 g L−1 glucose and 100 mg L−1 thiamine could be recommended for banana organogenesis. Results herein are of great importance and helpful in enhancing the commercial in vitro propagation protocols of banana, without the need of increasing the number of subcultures, which can cause somaclonal variation.


Banana (Musa acuminata Colla) Carbon source Cavendish Genetic fidelity Multiplication rate 



This work was conducted partially at Pomology and Genetics Departments, Faculty of Agriculture, Assiut University, Egypt.

Funding information

This work was financially supported by Assiut University.


  1. Abrahamian P, Kantharajah A (2011) Effect of vitamins on in vitro organogenesis of plant. Am J Plant Sci 2:669–674CrossRefGoogle Scholar
  2. Ahmad T, Abbasi NA, Hafiz IA, Ali A (2007) Comparison of sucrose and sorbitol as main carbon energy sources in micropropagation of peach rootstock GF-677. Pak J Bot 39:1269–1275Google Scholar
  3. Al-Khayri JM (2001) Optimization of biotin and thiamine requirements for somatic embryogenesis of date palm (Phoenix dactylifera L.). In Vitro Cell Dev Biol – Plant 37:453–456. CrossRefGoogle Scholar
  4. Al-Qurainy F, Nadeem M, Khan S, Alansi S, Tarroum M, Al-Ameri AA, Gaafar AZ, Alshameri A (2018) Rapid plant regeneration, validation of genetic integrity by ISSR markers and conservation of Reseda pentagyna an endemic plant growing in Saudi Arabia. Saudi J Biol Sci 25:111–116. CrossRefPubMedGoogle Scholar
  5. Bahmani R, Gholami M, Abdollahi H, Omid K (2009) The effect of carbon source and concentration on in vitro shoot proliferation of MM.106 apple rootstock. Fruit Veg Cereal Sci Biotech 3:35–37Google Scholar
  6. Baskaran P, Jayabalan N (2005) Role of basal media, carbon sources and growth regulators in micropropagation of Eclipta alba a valuable medicinal herb. KMITL Sci J 5:469–482Google Scholar
  7. Bohra P, Waman AA, Sathyanarayana BN, Umesha K, Gowda B (2016) Influence of different growth regulators on in vitro multiplication of mixed diploid banana (Musa AB). Proc Natl Acad Sci India Section B: Biol Sci 86:179–185CrossRefGoogle Scholar
  8. Brown PH, Hu H (1996) Phloem mobility of boron is species dependent: evidence for phloem mobility in sorbitol-rich species. Ann Bot 77:497–506CrossRefGoogle Scholar
  9. Buah JN, Kawamitsu Y, Yonemori S, Hayashi M, Murayama S (2000) Effects of various carbon sources and their combinations on in vitro growth and photosynthesis of banana plantlets. Plant Prod Sci 3:392–397CrossRefGoogle Scholar
  10. Castillo C, Fuller DQ (2016) Bananas: the spread of a tropical fruit as an agricultural staple. In: Lee-Thorp J, Katzenberg MA (eds) The Oxford handbook of the archaelogy of diet. Oxford University Press, Oxford. CrossRefGoogle Scholar
  11. Chowdhury MMH, Ashrafuzzaman M, Begum SN, Islam MM, Dhar P (2012) Regeneration of plantlets from grape (Vitis vinifera L.) through different explants. Int J Sustain Crop Prod 7:12–18Google Scholar
  12. Cuenca B, Vieitez AM (2000) Influence of carbon source on shoot multiplication and adventitious bud regeneration in in vitro beech cultures. Plant Growth Regul 32:1–12CrossRefGoogle Scholar
  13. De Paiva Neto VB, Otoni WC (2003) Carbon sources and their osmotic potential in plant tissue culture: does it matter? Sci Hortic 97:193–202CrossRefGoogle Scholar
  14. Ebadi M, Eghbali M (2017) The comparison of ISSR and RAPD markers with different species of Triticum. Biosci Biotechnol Res Commun 1:288–292Google Scholar
  15. Fuentes SR, Calheiros MB, Manetti-Filho J, Vieira LG (2000) The effects of silver nitrate and different carbohydrate sources on somatic embryogenesis in Coffea canephora. Plant Cell Tissue Organ Cult 60:5–13CrossRefGoogle Scholar
  16. George EF, Hall MA, De Klerk GJ (2008) The components of plant tissue culture media II: organic additions, osmotic and pH effects, and support systems. In: George EF, Hall MA, De Klerk GJ (eds) Plant propagation by tissue culture pp. Springer, Dordrecht, pp 115–173Google Scholar
  17. Goyer A (2010) Thiamine in plants: aspects of its metabolism and functions. Phytochemistry 71:1615–1624PubMedCrossRefGoogle Scholar
  18. Gupta M, Chyi YS, Romero-Severson J, Owen JL (1994) Amplification of DNA markers from evolutionarily diverse genomes using single primers of simple-sequence repeats. Theor Appl Genet 89:998–1006PubMedCrossRefGoogle Scholar
  19. Hossain MJ, Bari MA, Ara NA, Islam SS (2009) Effect of carbon sources on cell growth and regeneration ability in three cultivars of banana. J Biosci 17:83–88Google Scholar
  20. Hussein N (2012) Effects of nutrient media constituents on growth and development of banana (Musa spp.) shoot tips cultured in vitro. Afr J Biotechnol 11:9001–9006Google Scholar
  21. Ilczuk A, Jacygrad E (2016) In vitro propagation and assessment of genetic stability of acclimated plantlets of Cornus alba L. using RAPD and ISSR markers. In Vitro Cell Dev Biol – Plant 52:379–390PubMedPubMedCentralCrossRefGoogle Scholar
  22. Ill-Whan S, Karban SS (1998) Effects of media, carbon sources and cytokinins on shoot organogenesis in the Christmas tree, Scot pine (Pinus sylvestris). J Hortic Sci Biotechnol 73:822–827CrossRefGoogle Scholar
  23. Kadota M, Niimi Y (2004) Influences of carbon sources and their concentrations on shoot proliferation and rooting of Hosui’ Japanese pear. HortScience 39:1681–1683CrossRefGoogle Scholar
  24. Kumar U (2001) Methods in plant tissue culture. APH publishing corporation, New DelhiGoogle Scholar
  25. Lichtenthaler HK (1987) Chlorophylls and carotenoids: pigments of photosynthetic biomembranes. Methods Enzymol 148:350–382CrossRefGoogle Scholar
  26. Lucchesini M, Mensuali-Sodi A (2004) Influence of medium composition and vessel ventilation on in vitro propagation of Phillyrea latifolia L. Sci Hortic 100:117–125CrossRefGoogle Scholar
  27. Madhulatha P, Kirubakaran SI, Sakthivel N (2006) Effects of carbon sources and auxins on in vitro propagation of banana. Biol Plant 50:782–784CrossRefGoogle Scholar
  28. Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  29. Nissen O (1984) MSTAT. A microcomputer program for statistical analyses of experiments and surveys. In: H Riley, Skjelvag AO (eds) The impact of climate on grass production and quality. Proc General Meet Eur Grassl Fed, 10th, As, Norway. 26–30 June 1984. Norwegian state agric. Res. Stations, As, pp 555–559Google Scholar
  30. Prabhuling G, Sathyanarayana BN (2017) Cheaper carbon sources for micropropagation of banana cv. 'Grande Naine'. International J Agric Sci 13:124–131Google Scholar
  31. Preethi D, Sridhar TM, Naidu CV (2011) Carbohydrate concentration influences on in vitro plant regeneration in Stevia rebaudiana. J Phytol 3:61–64Google Scholar
  32. Qamar M, Qureshi ST, Khan IA, Raza S (2015) Optimization of in vitro multiplication for exotic banana (Musa spp.) in Pakistan. Afr J Biotechnol 14:1989–1995. CrossRefGoogle Scholar
  33. Reddy MP, Sarla N, Siddiq EA (2002) Inter simple sequence repeat (ISSR) polymorphism and its application in plant breeding. Euphytica 128:9–17CrossRefGoogle Scholar
  34. Redei GP (1974) ‘Fructose effect’ in higher plants. Ann Bot 38:287–297CrossRefGoogle Scholar
  35. Robinson JC (1996) Bananas and plantains. CAB international, Wallingford, p 238Google Scholar
  36. Rolland F, Moore B, Sheen J (2002) Sugar sensing and signaling in plants. Plant Cell 14:185–205CrossRefGoogle Scholar
  37. Rolland F, Baena-Gonzalez E, Sheen J (2006) Sugar sensing and signaling in plants: conserved and novel mechanisms. Annu Rev Plant Biol 57:675–709PubMedCrossRefPubMedCentralGoogle Scholar
  38. Romano A, Noronha C, Martins-Loucao MA (1995) Role of carbohydrates in micropropagation of cork oak. Plant Cell Tissue Organ Cult 40:159–167CrossRefGoogle Scholar
  39. Ruzic DV, Lazic TI, Cerovic RM (2008) Micropropagation of some Prunus and Pyrus genotypes in vitro as affected by different carbon sources. Acta Hortic 795:413–418. CrossRefGoogle Scholar
  40. Saha S, Adhikari S, Dey T, Ghosh P (2016) RAPD and ISSR based evaluation of genetic stability of micropropagated plantlets of Morus alba L. variety S-1. Meta Gene 7:7–15. CrossRefPubMedPubMedCentralGoogle Scholar
  41. Singh MP, Kumar S (2009) Plant tissue culture. APH Publishing, New DelhiGoogle Scholar
  42. Skylar A, Sung F, Hong F, Chory J, Wu X (2011) Metabolic sugar signal promotes Arabidopsis meristematic proliferation via G2. Dev Biol 351:82–89PubMedCrossRefPubMedCentralGoogle Scholar
  43. Sotiropoulos TE, Molassiotis AN, Mouhtaridou GI, Papadakis I, Dimassi KN, Therios IN, Diamantidis G (2006) Sucrose and sorbitol effects on shoot growth and proliferation in vitro, nutritional status and peroxidase and catalaze isoenzymes of M 9 and MM106 apple (Malus domestica Borkh.) rootstocks. Eur J Hortic Sci 71:114–119Google Scholar
  44. Ssekiwoko F, Talengera D, Kiggundu A, Namutebi MK, Karamura E, Kunert K (2014) In-vitro proliferation of Musa balbisiana improves with increased vitamin concentration and dark culturing. J Appl Biol Biotechnol 2:1–7Google Scholar
  45. Strosse H, Domergue R, Panis B, Escalant J-V, Côte F (2003) Banana and plantain embryogenic cell suspensions. In: Vézina A, Picq C (eds) INIBAP technical guidelines. The international network for the improvement of Banana and plantain, Montpellier, p 8Google Scholar
  46. Vasconcelos JM, Saldanha CW, Dias LLC, Maldaner J, Rêgo MM, Silva LC, Otoni WC (2014) In vitro propagation of Brazilian ginseng [Pfaffia glomerata (Spreng.) Pedersen] as affected by carbon sources. In Vitro Cell Dev Biol – Plant 50:746–751CrossRefGoogle Scholar
  47. Venkatachalam L, Sreedhar RV, Bhagyalakshmi N (2007a) Micropropagation in banana using high levels of cytokinins does not involve any genetic changes as revealed by RAPD and ISSR markers. Plant Growth Regul 51:193–205. CrossRefGoogle Scholar
  48. Venkatachalam L, Sreedhar RV, Bhagyalakshmi N (2007b) Molecular analysis of genetic stability in long-term micropropagated shoots of banana using RAPD and ISSR markers. Electron J Biotechnol.
  49. Vora NC, Jasrai YT (2011) Effect of various carbon sources on in vitro shoot multiplication of banana. Phytomorphology 61:111–116Google Scholar
  50. Waman AA, Bohra P, Sathyanarayana BN (2014) Not all sugars are sweet for banana multiplication; in vitro multiplication, rooting and acclimatization of banana as influenced by carbon sources concentrations interactions. In Vitro Cell Dev Biol – Plant 50:552–560CrossRefGoogle Scholar
  51. Waman AA, Bohra P, Sathyanarayana BN (2016) Effect of various cytokinins and auxins on in vitro regeneration of plantlets from isolated bud clumps of silk Banana var. Nanjanagud Rasabale (Musa AAB). Indian J Plant Physiol 21:64–69CrossRefGoogle Scholar
  52. Williams JGK, Kubelik AR, Livak KJ, Rafalski JA, Tingey SV (1990) DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic Acids Res 18:6531–6535PubMedPubMedCentralCrossRefGoogle Scholar
  53. Youssef M, James A, Mayo-Mosqueda A, Ku-Cauich JR, Grijalva-Arango R, Escobedo-GM RM (2010) Influence of the genotype and age of the explant source on the capacity for somatic embryogenesis of two Cavendish banana cultivars (Musa acuminata Colla, AAA). Afr J Biotechnol 9:2216–2223Google Scholar
  54. Youssef M, Valdez-Ojeda R, Ku-Cauich JR, Escobedo-GraciaMedrano RM (2015) Enhanced protocol for isolation of plant genomic DNA. J Agri Environ Sci 4:172–180. CrossRefGoogle Scholar

Copyright information

© The Society for In Vitro Biology 2019

Authors and Affiliations

  1. 1.Pomology Department, Faculty of AgricultureAssiut UniversityAssiutEgypt
  2. 2.Genetics Department, Faculty of AgricultureAssiut UniversityAssiutEgypt

Personalised recommendations