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Highly efficient shoot regeneration of Bacopa monnieri (L.) using a two-stage culture procedure and assessment of genetic integrity of micropropagated plants by RAPD

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Abstract

A two-stage culture procedure has been developed for highly efficient shoot regeneration from leaf and internode explants of Bacopa monnieri. Adventitious shoot buds were obtained on the shoot induction medium containing Murashige and Skoog’s (MS) basal salt supplemented with 1.5 mg/l thidiazuron and 0.5 mg/l naphthalene acetic acid; these shoot buds were subcultured on the multiplication (second) medium amended with BAP (benzyl amino purine). Multiplication medium containing 0.5 mg/l BAP produced more shoots (135) and longer shoots (7.8 cm) with more nodes (6). Best response of root induction with more number of roots (16.5) and longer roots (8.7 cm) was observed in half strength MS basal medium supplemented with 1.0 mg/l IBA (indole-3-butyric acid) and 0.5 mg/l phloroglucinol. In vitro obtained plants were transferred to the field after hardening with a 100% survival rate. Random amplified polymorphic DNA analysis was carried out using five random primers. The amplification products were monomorphic in micropropagated plants and similar to those of mother plant. No polymorphism was detected revealing the genetic integrity of micropropagated plants.

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Abbreviations

BAP:

N6-Benzyl amino purine

CA:

Chlorogenic acid

IBA:

Indole-3-butyric acid

IAA:

Indole-3-acetic acid

Kin:

Kinetin

MS:

Murashige and Skoog’s (1962)

NAA:

Naphthalene acetic acid

PG:

Phloroglucinol

PGR:

Plant growth regulator

RAPD:

Random amplified polymorphic DNA

SA:

Salicylic acid

TDZ:

Thidiazuron

References

  • Ali G, Shrivastav PS, Iabal M (2001) Responses of Bacopa monnera cultures to cadmium toxicity. Bull Environ Con Tox 66:342–349

    Article  CAS  Google Scholar 

  • Anonoymous (1997) Indian medicinal plants: a sector study. Occasional paper No. 54. Export-Import Bank of India, Quest publications, Bombay, India

  • Capelle S, Mok D, Kirchner S, Mok S (1983) Effects of thidiazuran on (Δ2-isopentyl) (8–14C) adenosine in callus tissues of phaseolus linatus L. Plant Physiol 73:796–802

    Article  CAS  PubMed  Google Scholar 

  • David JJ (1983) Adventitious root formation ‘in vitro’ in apple rootstocks (Malus pumila) I. Factors affecting the length of the auxin-sensitive phase in M9. Physiol Plant 5:149–153

    Google Scholar 

  • De Klerk GJ, Vander Kreiken W, De Jong JC (1999) The formation of adventitious roots: new concepts, new possibilities. In Vitro Cell Dev Biol Plant 35:189–199

    Article  Google Scholar 

  • Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation. Version II. Plant Mol Biol Rep 1:19–21

    Article  CAS  Google Scholar 

  • Dimitrov B, Tashera K, Zagorska N, Evstatiera L (2003) In vitro cultivation of Rhodiola rosea L. Gene Breed 32:3–6

    Google Scholar 

  • Faisal M, Anis M (2005) An efficient in vitro method for mass propagation of Tylophora indicia. Biol Plant 49:257–260

    Article  Google Scholar 

  • Geetha N, Venkatachalam P, Prakash V, Lakshmi Sita G (1998) High frequency induction of multiple shoots and plant regeneration from seedling explants of pigeonpea (Cajanus cajan L.). Curr Sci 75:1036–1041

    CAS  Google Scholar 

  • Giridhar P, Gururaj H, Ravishankar G (2005) In vitro shoot multiplication through shoot tip cultures of Decalepis hamiltonii Wight & Arn., a threatened plant endemic to Southern India. In Vitro Cell Dev Biol Plant 41:77–80

    Google Scholar 

  • González A, Casares TR, Sánchez RodríguezR (1991) Adventitious root induction in Corylusvellana L. cotyledon. In Vitro Cell Dev Bio Plant 27:125–131

    Article  Google Scholar 

  • Hoque A, Arima S (2002) Overcoming phenolic accumulation during callus induction and in vitro organogenesis in water chestnut (Trapa japonica Flerov.). In Vitro Cell Dev Biol Plant 38:342–346

    Article  CAS  Google Scholar 

  • Husain MK, Avis M, Shahzad A (2007) In vitro propagation of Indian kino (Pterocarpus marsupium. Rokb.) using Thidiazuron. In Vitro Cell Dev Biol Plant 43:59–64

    CAS  Google Scholar 

  • Ibrahim R, Debergh PC (2001) Factors controlling high efficiency adventitious bud formation and plant regeneration from in vitro leaf explants of roses (Rosa hybrida L.). Sci Hortic 88:41–57

    Article  Google Scholar 

  • Iram S, Anis M (2007) In vitro shoot multiplication and plantlet regeneration from nodal explants of Cassia angustifolia (Vahl.), a medicinal plant. Acta Phys Plant 29:233–238

    Article  Google Scholar 

  • Jayanthi M, Mandal PK (2001) Plant regeneration through somatic embryogenesis and RAPD analysis of regenerated plants in Tylophora indica (Burm. F. Merrill.). In Vitro Cell Dev Biol Plant 37:576–580

    Article  CAS  Google Scholar 

  • Kanita A, Kothari SL (2002) High efficiency adventious shoot bud formation and plant regeneration from leaf explants of Dianthus chinesis L. Sci Hortic 90:205–212

    Google Scholar 

  • Kawiak AA, Lojkowska E (2004) Application of RAPD in the determination of genetic fidelity in micropropagated drosera plantlets in vitro cell. Dev Biol Plant 40:592–595

    CAS  Google Scholar 

  • Konan NK, Sangwan RS, Sangwan-Norreel BS (1994) Efficient in vitro shoot-regeneration systems in Cassava (Manihot esculenta Crantz). Plant Breed 113:227–236

    Article  Google Scholar 

  • Liu HJ, Xu YAN, Liu Y-J, Liu C-Z (2006) Plant regeneration from leaf explants of Rhodiola fastigiata. In Vitro Cell Dev Biol Plant 42:345–347

    Article  CAS  Google Scholar 

  • Magioli C, Rocha AP, Oiveira DE (2004) Efficient shoot organogenesis of eggplant (Solanum melongena L.) induced by thidiazuron. Plant Cell Rep 7:661–663

    Google Scholar 

  • Mok MC, Mok DW, Amstrong DJ, Shudo K, Isogai Y, Okamanto T (2005) Cytokinin activity of N-phenyl-N′-1, 2, 3-thiadiazol-5-urea (thidiazuron). Phytochemistry 21:1509–1511

    Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and biossays with tobacco tissue cultures. Physiol Plant 15:497–573

    Article  Google Scholar 

  • Prakash S, Elangomathavan R, Seshadri S, Kathiravan K, Ignacimuthu S (2004) Efficient regeneration of Curcuma amada Roxb plantlets from rhizome and leaf sheath explants. Plant Cell Tissue Organ Cult 78:159–165

    Article  CAS  Google Scholar 

  • Rahman LU, Verma PC, Singh D, Gupta MM, Banerjee S (2002) Bacosides production by suspension cultures of Bacopa monnieri (L). Biotechnol Lett 24:1427–1429

    Article  Google Scholar 

  • Ramana M, Gosukonda, Ananta Porobodessai, Essie Blay, Channapatna S, Prakasha Curt M, Peterson (1995) Thidiazuron-induced adventitious shoot regeneration of sweet potato (Ipomoea batatas). In Vitro Cell Dev Biol Plant 31:65–71

  • Rout GR, Das (2002) An assessment of genetic integrity of micropropagated plants of Plumbago zeylanica by RAPD markers. Biol Plant 45:27–32

  • Rout GR, Das P, Goel S, Raina SN (1998) Determination of genetic stability of micropropagated plants of ginger using random amplified polymorphic DNA (RAPD) markers. Bot Bull Acad Sin 39:23–27

    CAS  Google Scholar 

  • Selvi DN, George L, Eapen S (2002) Micropropagation and field evaluation of micropropagated plants of turmeric. Plant Cell Tissue Organ Cult 68:143–151

    Article  Google Scholar 

  • Sen J, Sen S (1995) Two-step bud culture technique for a high frequency regeneration of Gladiolus corms. Sci Hortic 64:133–138

    Article  Google Scholar 

  • Shrivastav N, Ranjani M (1999) Multiple shoot regeneration and tissue culture studies on Bacopa monnieri (L.) Pennel. Plant Cell Rep 18:919–923

    Article  Google Scholar 

  • Shu QY, Liu GS, Qi DM, Chu CC, Liu J, Li HJ (2003) An effective method for axillary bud culture and RAPD analysis of cloned plants in tetraploid black locust. Plant Cell Rep 22:175–180

    Article  CAS  PubMed  Google Scholar 

  • Shukla OP, Pubey S, Ravi VN (2007) Preferential accumulation of cadmium and chromium: toxicity in Bacopa monnieri. L. under mixed metal treatments. Bull Environ Contam Toxicol 78:252–257

    Article  CAS  PubMed  Google Scholar 

  • Singh HK, Rastogi RP, Sriman RC, Dhawan BN (1988) Effect of bacoside A and B on avoidance response in rats. Phytother Res 2:70–75

    Article  CAS  Google Scholar 

  • Skala E, Wysokinska H (2004) In vitro regeneration of Salvia nemorosa L. from shoot tips and leaf explants. In Vitro Cell Dev Biol Plant 40:596–602

    Article  CAS  Google Scholar 

  • Stough C, Lloyd J, Clarke J, Downey LA, Hutchison CW, Rodgers T, Nathan PJ (2001) The chronic effects of an extract Bacopa monniera (Brahimi) on cognitive function in healthy human subjects. Psychopharmacology 156:481–484

    Article  CAS  PubMed  Google Scholar 

  • Tang W, Guo Z (2001) In vitro propagation of loblolly pine via direct somatic organogenesis from mature cotyledons and hypocotyls. Plant Growth Regul 33:25–31

    Article  Google Scholar 

  • Thomas JC, Katterman FR (1986) Cytokinin activity induced by thidiazuran. Plant Physiol 81:681–683

    Article  CAS  PubMed  Google Scholar 

  • Tiwari V, Deo Singh B, Nath Tiwari K (1998) Shoot regeneration and Somatic embryogenesis from different explants of Brahmi (Bacopa monniera (L.) Wettst.). Plant Cell Rep 17:538–543

    Article  CAS  Google Scholar 

  • Tiwari V, Tewari KN, Singh BD (2000) Suitability of liquid cultures for in vitro multiplication of Bacopa Monniera (L.) Wettst. Phytomorphology 50:337–342

    Google Scholar 

  • Tiwari V, Tewari KN, Singh BD (2001) Comparative studies of cytokinin on in vitro propagation of Bacopa monniera. Plant Cell Tissue Organ Cult 66:9–16

    Article  CAS  Google Scholar 

  • Tiwari V, Tewari KN, Singh BD (2006) Shoot bud regeneration from different explants of Bacopa monniera (L.) Wettst. by trimethoprim and bavistin. Plant Cell Rep 25:629–635

    Article  CAS  PubMed  Google Scholar 

  • Venkatachalam LE, Sreedhar RV, Bhagyalakshmi N (2007) 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

    Article  CAS  Google Scholar 

  • 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 18:6531–6535

    Article  CAS  PubMed  Google Scholar 

  • Zanol GC, Fortes GR, de L, Campos AD, Centellas AQ, da Silva JB (1998) In vitro rooting and peroxidase activity of apple rootstock treated with indole butyric acid and phloroglucinol. Res Brasil Fisiol Veg 10:65–68

    CAS  Google Scholar 

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Authors and Affiliations

  1. Entomology Research Institute, Loyola College, Chennai, 600034, Tamilnadu, India

    S. Antony Ceasar & Savarimuthu Ignacimuthu

  2. Department of Biotechnology, St. Peter’s Engineering College, Avadi, Chennai, 600054, India

    S. Antony Ceasar, S. Lenin Maxwell, K. Bhargav Prasad & M. Karthigan

Authors
  1. S. Antony Ceasar
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  2. S. Lenin Maxwell
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  3. K. Bhargav Prasad
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  4. M. Karthigan
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  5. Savarimuthu Ignacimuthu
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Correspondence to Savarimuthu Ignacimuthu.

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Communicated by E. Lojkowska.

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Antony Ceasar, S., Lenin Maxwell, S., Bhargav Prasad, K. et al. Highly efficient shoot regeneration of Bacopa monnieri (L.) using a two-stage culture procedure and assessment of genetic integrity of micropropagated plants by RAPD. Acta Physiol Plant 32, 443–452 (2010). https://doi.org/10.1007/s11738-009-0419-8

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  • DOI: https://doi.org/10.1007/s11738-009-0419-8

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