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In vitro regeneration and Agrobacterium tumefaciens-mediated genetic transformation of Parkia timoriana (DC.) Merr.: a multipurpose tree legume

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Abstract

In vitro regeneration of Parkia timoriana (DC.) Merr. has been achieved using cotyledonary node explants. The ability to produce multiple shoots has been evaluated using semi-solid Murashige and Skoog (MS) basal medium and Gamborg’s B-5 basal medium supplemented with various concentrations of α-naphthalene acetic acid (NAA) and 6-benzylaminopurine (BA) either in single or in combinations. The explants cultured in MS medium supplemented with combinations of 2.7 μM NAA and 11 μM BA showed the maximum frequency of multiple shoots (96.66%) formation and number of shoots per explants (6.60), respectively. For rooting, full and half strength MS medium supplemented with various concentrations of indole-3-butyric acid (IBA) and NAA were studied and the highest number of root formation was observed in full-strength MS supplemented with 9.8 μM IBA. Using Agrobacterium tumefaciens strain EHA105 pCAMBIA2301 various optimum conditions for efficient transformation were determined by recording the percentage of GUS+ explants. Following the optimized conditions, the co-cultured explants were cultured on semi-solid shoot regeneration medium containing MS medium + 2.7 μM NAA + 11 μM BA + 100 mg/l kanamycin + 500 mg/l cefotaxime. After 8 weeks of culture, the regenerated shoots were rooted in rooting medium (RM) containing MS medium + 9.8 μM indole-3-butyric acid (IBA), 3% sucrose, 7.5 mg/l kanamycin and 500 mg/l cefotaxime. Successful transformation was confirmed by histochemical GUS activity of the regenerated shoots, nptII gene PCR analyses of the regenerated kanamycin resistant plantlets and Southern analysis of putative transgenic PCR+ plants.

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Abbreviations

MS:

Murashige and Skoog (1962)

B-5:

Gamborg et al. (1968)

NAA:

α-Naphthalene acetic acid

BA:

6-Benzylaminopurine

IBA:

Indole-3-butyric acid

GUS:

β-Glucuronidase

CaMV:

35S 35S promoter of the cauliflower mosaic virus

nptII:

Neomycin phosphotransferase

PCR:

Polymerase chain reaction

OD600 :

Optical density at 600 nm

References

  • Ajithkumar D, Seeni S (1998) Rapid cloncal multiplication through in vitro axillary shoot proliferation of Aegle marmelos (L.) Corr. a medicinal tree. Plant Cell Rep 17:422–426. doi:10.1007/s002990050418

    Article  CAS  Google Scholar 

  • Al-Wasel AS (2000) Micropropagation of Acacia seyal Del. in vitro. J Arid Environ 46:425–431. doi:10.1006/jare.2000.0687

    Article  Google Scholar 

  • Amoo SO, Ayisire BE (2005) Induction of callus and somatic embryogenesis from cotyledon explants of Parkia biglobosa (Jacq.) Benth. Afr J Biotechnol 4:68–71

    CAS  Google Scholar 

  • Arumugam S, Rao MV (1996) In vitro production of plantlets from cotyledonary node cultures of Aegle marmelos (L.) Corr. Adv Plant Sci 9:181–186

    Google Scholar 

  • Baulcombe D (2004) RNA silencing in plants. Nature 431:356–363

    Article  PubMed  CAS  Google Scholar 

  • Cervera M, Ortega C, Navarro A, Navarro L, Pena L (2000) Generation of transgenic citrus plants with tolerance-to-salinity gene HAL2 from yeast. J Hortic Sci Biotechnol 75:26–30

    CAS  Google Scholar 

  • Chilton MD, Currier TC, Farrand SK, Bendich AJ, Gordon MP, Nestor EW (1974) Agrobacterium tumefaciens DNA and PS8 bacteriophage DNA not detected in crown gall tumors. Proc Natl Acad Sci USA 71:3672–3676

    Article  PubMed  CAS  Google Scholar 

  • Datta SK, Datta K (1983) Auxin induced regeneration of forest tree—Dalbergia sissoo Roxb. through tissue culture. Curr Sci 52:434–436

    CAS  Google Scholar 

  • Dewan A, Nanda K, Gupta SC (1992) In vitro micropropagation of Acacia nilotica sub sp indica Brenan via cotyledonary nodes. Plant Cell Rep 12:18–21. doi:10.1007/BF00232415

    Article  Google Scholar 

  • Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements for superior cultures of soybean root cells. Exp Cell Res 50:151–158. doi:10.1016/0014-4827(68)90403-5

    Article  PubMed  CAS  Google Scholar 

  • Hopkins HCF (1994) The Indo-Pacific species of Parkia (Leguminosae: Mimosoideae). Kew Bull 49:181–234

    Article  Google Scholar 

  • Hossain M, Islam R, Karim MR, Joarder OI, Biswas BK (1994) Regeneration of plantlets from in vitro cultured cotyledons of Aegle marmelos Corr (Rutaceae). Sci Hort 57:315–321. doi:10.1016/0304-4238(94)90114-7

    Article  Google Scholar 

  • Jefferson RA (1987) Assaying chimeric in plants: the GUS gene fusion system. Plant Mol Bio Rep 5:387–405. doi:10.1007/BF02667740

    Article  CAS  Google Scholar 

  • Jaiwal PK, Kumari R, Ignacimuthu S, Potrykus I, Sautter C (2001) Agrobacterium tumfefaciens-mediated genetic transformation of mungbean (Vigna radiata L. Wilczek)—a recalcitrant grain legume. Plant Sci 161:239–247. doi:10.1016/S0168-9452(01)00352-1

    Article  PubMed  CAS  Google Scholar 

  • Joshi I, Bisht P, Sharma VK, Uniyal DP (2003) Studies on effect of nutrient media for clonal propagation of superior phenotypes of Dalbergia sissoo Roxb. through tissue culture. Silvae Genetica 52:3–4

    Google Scholar 

  • Kanjilal UN, Kanjilal PC, Das A (1982) Flora of Assam, vol 2. Avon, Delhi, p 151

  • Karthikeyan AS, Sarma KS, Veluthambi K (1996) Agrobacterium tumefaciens-mediated transformation of Vigna mungo L. Hepper. Plant Cell Rep 15:328–331. doi:10.1007/BF00232365

    Article  CAS  Google Scholar 

  • Longvah T, Deosthale YG (1998) Nutrient composition and food potential of Parkia roxburghii, a less known tree legume from northeast India. Food Chem 62:477–481. doi:10.1016/S0308-8146(97)00179-9

    Article  CAS  Google Scholar 

  • Mittal A, Agarwal R, Gupta SC (1989) In vitro developments of plantlets from axillary buds of Acacia auriculiformis—a leguminous tree. Plant Cell Tissue Org Cult 19:55–60. doi:10.1007/BF00037777

    Article  Google Scholar 

  • Moore GA, Jacomo CC, Neidigh JL, Lawrence SD, Cline K (1992) Agrobacterium-mediated transformation of Citrus stem segments and regeneration of transgenic plants. Plant Cell Rep 11:238–242. doi:10.1007/BF00235073

    Article  CAS  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol Plant 15:473–491. doi:10.1111/j.1399-3054.1962.tb08052.x

    Article  CAS  Google Scholar 

  • Pradhan C, Kar S, Pattnaik S, Chand PK (1998) Propagation of Dalbergia sissoo Roxb through in vitro shoot proliferation from cotyledonary nodes. Plant Cell Rep 18:122–126. doi:10.1007/s002990050543

    Article  CAS  Google Scholar 

  • Perez-Molphe-Balch E, Ochoa-Alejo N (1998) Regeneration of transgenic plants of Mexican lime from Agrobacterium rhizogenes-transformed tissues. Plant Cell Rep 17:591–596. doi:10.1007/s002990050448

    Article  CAS  Google Scholar 

  • Purohit SD, Dave A (1996) Micropropagation of Sterculia urens Roxb an endangered tree species. Plant Cell Rep 15:704–706. doi:10.1007/BF00231929

    Article  CAS  Google Scholar 

  • Saklani A, Rao RR (2002) In: Rao (ed.) Advances in legume research in India. Bishen Singh Mahendra Pal Singh, Dehra Dun, pp 239–250

    Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  • Sharma A, Haridasan K, Borthakur K (2002) In: Rao (ed.) Advances in legume research in India. Bishen Singh Mahendra Pal Singh, Dehra Dun, pp 171–179

    Google Scholar 

  • Suvachittanont W, Kurashima Y, Esumi H, Tsuda M (1996) Formation of thiazolidine-4-carboxylic acid (thioproline), an effective nitrite-trapping agent in human body, in Parkia speciosa seeds and other edible leguminous seeds in Thailand. Food Chem 55:359–363. doi:10.1016/0308-8146(95)00132-8

    Article  CAS  Google Scholar 

  • Tahira T, Tsuda M, Wakabayashi K, Nagao M, Sugumura T (1984) Kinetics of nitrosation of a major nitroso compound in human urine and its role as nitrite scavenger. Gann 75:889–894

    PubMed  CAS  Google Scholar 

  • Tahira T, Ohgaki H, Wakabayashi K, Nagao M, Sugumura T (1988) Inhibitory effect of thioproline on carcinogenesis induced by N-benzylmethylamine and nitrite. Food Chem Toxicol 26:511–518. doi:10.1016/0278-6915(88)90003-8

    Article  PubMed  CAS  Google Scholar 

  • Thangjam R, Damayanti M, Jitendra GS (2003a) Cadra cautella Walker (Lepidoptera: Crambidae: Phycitinae)—a pest on Parkia timoriana (DC.) Merr. in Manipur. Curr Sci 85:725–726

    Google Scholar 

  • Thangjam R, Damayanti M, Jitendra GS (2003b) A simple and rapid method for isolation of DNA from imbibed embryos of Parkia timoriana (DC.) Merr. for PCR analysis. J Food Agric Environ 1:36–38

    CAS  Google Scholar 

  • Thangjam R, Maibam RS (2006) Induction of callus and somatic embryogenesis of cotyledonary explants of Parkia timoriana (DC.) Merr., a multipurpose tree legume. J Food Agric Environ 4:335–339

    Google Scholar 

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Authors and Affiliations

  1. Department of Biotechnology, School of Life Sciences, Mizoram University, Aizawl, 796004, Mizoram, India

    Robert Thangjam

  2. Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, 781039, Assam, India

    Lingaraj Sahoo

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  1. Robert Thangjam
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  2. Lingaraj Sahoo
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Correspondence to Robert Thangjam.

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Communicated by T. Moriguchi.

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Thangjam, R., Sahoo, L. In vitro regeneration and Agrobacterium tumefaciens-mediated genetic transformation of Parkia timoriana (DC.) Merr.: a multipurpose tree legume. Acta Physiol Plant 34, 1207–1215 (2012). https://doi.org/10.1007/s11738-011-0917-3

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  • DOI: https://doi.org/10.1007/s11738-011-0917-3

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