SpringerLink
Log in

Genetic transformation of Indian isolate of Lemna minor mediated by Agrobacterium tumefaciens and recovery of transgenic plants

  • Research Article
  • Published:
Physiology and Molecular Biology of Plants Aims and scope Submit manuscript

Abstract

Transgenic plants of an Indian isolate of Lemna minor have been developed for the first time using Agrobacterium tumefaciens and hard nodular cell masses ‘nodular calli’ developed on the BAP - pretreated daughter frond explants in B5 medium containing sucrose (1.0 %) with 2,4-D (5.0 μM) and 2-iP (50.0 μM) or 2,4-D (50.0 μM) and TDZ (5.0 μM) under light conditions. These calli were co-cultured with A. tumefaciens strain EHA105 harboring a binary vector that contained genes for β-glucuronidase with intron and neomycin phosphortransferase. Transformed cells selected on kanamycin selection medium were regenerated into fronds whose transgenic nature was confirmed by histochemical assay for GUS activity, PCR analysis and Southern hybridization. The frequency of transformation obtained was 3.8 % and a period of 11–13 weeks was required from initiation of cultures from explants to fully grown transgenic fronds. The pretreatment of daughter fronds with BAP, use of non-ionic surfactant, presence of acetosyringone in co-cultivation medium, co-culture duration of 3 d and 16 h photoperiod during culture were found crucial for callus induction, frond regeneration and transformation of L. minor. This transformation system can be used for the production of pharmaceutically important protein and in bioremediation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Abbreviations

2,4-D:

2,4-dichlorophenoxyacetic acid

2-iP:

6-(-γ,-γ-dimethyl allyl amino) purine

BAP:

6-benzylaminopurine

IAA:

Indole-3-acetic acid

Kin:

Kinetin

TDZ:

Thidiazuron

CaMV:

Cauliflower mosaic virus

References

  • Bog M, Baumbach H, Schween U, Hellwig F, Landolt E, Appenroth KJ (2010) Genetic structure of the genus Lemna L. (Lemnaceae) as revealed by amplified fragment length polymorphism. Planta 232:609–619

    Article  PubMed  CAS  Google Scholar 

  • Brain RA, Solomon KR (2007) A protocol for conducting 7-day daily renewal tests with Lemna gibba. Nat Prot 2:979–987

    Article  CAS  Google Scholar 

  • Brunning JL, Kintz BL (1977) Computational hand book of statistics, 2nd edn. Scott Foresman, Glenview

    Google Scholar 

  • Cabrera LI, Salazar GA, Chase MW, Mayo SJ, Bogner J, Davila P (2008) Phylogenetic relationships of aroids and duckweeds (Araceae) inferred from coding and noncoding plastid DNA. Am J Bot 95:1153–1165

    Article  CAS  Google Scholar 

  • Cheng JJ, Stomp AM (2009) Growing duckweed to recover nutrients from wastewaters and for production of fuel ethanol and animal feed. Clean 37:17–26

    CAS  Google Scholar 

  • Cheng M, Fry JE, Pang S, Zhou H, Hironaka CM, Duncan DR, Conner TW, Wan Y (1997) Genetic transformation of wheat mediated by Agrobacterium tumefaciens. Plant Physiol 115:971–980

    PubMed  CAS  Google Scholar 

  • Chhabra G, Singh RP, Jaiwal PK (2007) Duckweed biotechnology for commercial exploitation. Physiol Mol Biol Plant 13:1–7

    CAS  Google Scholar 

  • Chung MH, Chen MK, Pan SM (2000) Floral spray transformation can efficiently generate Arabidopsis transgenic plants. Transgen Res 9:471–476

    Article  CAS  Google Scholar 

  • Cox KM, Sterling JD, Regan JT, Gasdaska JR, Frantz KK, Peele CG, Black A, Passmore D, Moldovan-Loomis C, Srinivasan M, Cuison S, Cardarelli PM, Dickey LF (2006) Glycan optimization of a human monoclonal antibody in the aquatic plant Lemna minor. Nat Biotech 24:1591–1597

    Article  CAS  Google Scholar 

  • Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements of suspension cultures of soybean root cells. Exp Cell Res 50:151–156

    Article  PubMed  CAS  Google Scholar 

  • Hagner M, Penttinen O, Pasanen T, Tiilikkala K, Setala H (2010) Acute toxicity of birch tar oil on aquatic organisms. Agri Food Sci 19:24–33

    Article  Google Scholar 

  • Haustein AT, Gilman RH, Skillicorn PW, Ventura G (1987) Safety and efficacy of sewage grown Lemna as a protein source for chickens. Sunny Spring, Columbia

    Google Scholar 

  • Jefferson RA, Kavanagh TA, Beven MW (1987) GUS fusions: betaglucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907

    PubMed  CAS  Google Scholar 

  • Khellaf N, Zerdaoui M (2009) Growth response of the duckweed Lemna minor to heavy metal pollution. Iran J Environ Health Sci Engg 6:161–166

    CAS  Google Scholar 

  • Kim MJ, Baek K, Park CM (2009) Optimization of conditions for transient Agrobacterium-mediated gene expression assays in Arabidopsis. Plant Cell Rep 28:1159–1167

    Article  PubMed  CAS  Google Scholar 

  • Kruse C, Boehm R, Voeste D (2002) Transient transformation of Wolffia columbiana by particle bombardment. Aquat Bot 72:175–181

    Article  CAS  Google Scholar 

  • Landolt E (1986) The family of Lemnaceae—a monographic study. Vol 1 Veröffentlichungen des Geobotanischen Institutes der ETH, Stiftung Rübel (Zürich) 71:1–566

    Google Scholar 

  • Leblebici Z, Aksoy A (2010) Growth and lead accumulation capacity of Lemna minor and Spirodela polyrhiza (Lemnaceae): Interactions with nutrient enrichment. Water Air Soil Poll. (published online) doi:10.1007/s11270-010-0413-1

  • Les DH, Crawford DJ, Landolt E, Gabel JD, Kimbell RT (2002) Phylogeny and systematics of Lemnaceae, the duckweed family. Syst Bot 27:221–240

    Google Scholar 

  • Li J, Jain M, Vunsh R, Vishnevetsky J, Hanania U, Flaishman M, Perl A, Edelman M (2004) Callus induction and regeneration in Spirodela and Lemna. Plant Cell Rep 22:457–464

    Article  PubMed  CAS  Google Scholar 

  • Men BX, Ogle RB, Preston TR (1995) Use of duckweed (Lemna spp.) as replacement for soybean meal in as basal diet of broken rice for fattening ducks. Livest Res Rur Dev 7:5–8

    Google Scholar 

  • Men LT, Van BH, Chinh MT, Preston TR (1997) Effect of dietary protein level and duckweed (Lemna spp.) on reproductive performance of pigs fed a diet of ensiled cassava root or cassava root meal. Livest Res Rur Dev 9:1–5

    Google Scholar 

  • OECD guidelines for the testing of chemicals (2002) Lemna sp. Growth inhibition test. Organization for Economic Cooperation and Development, Berlin, p 22

    Google Scholar 

  • Porath D (1993) Aquaculture production of duckweeds for human consumption. United States Kibbutz, Tzora (IL) 5269819. www.freepatentsonline.com/5269819.html accessed on August 17, 2010

  • Roger SO, Bendich AJ (1988) Extraction of DNA from plant tissues. In: Gelvin SB, Schilperoot RA (eds) Plant molecular biology manual. Kluwer, Dordrecht, pp 1–11

    Google Scholar 

  • Saini R, Jaiwal PK (2007) Agrobacterium tumefaciens—mediated transformation of blackgram: an assessment of factors influencing the efficiency of uidA gene transfer. Biol Plant 51:69–79

    Article  CAS  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, New York

    Google Scholar 

  • Skillicorn P, Spira W, Journey W (1993) Duckweed aquaculture—a new aquatic farming system for develo** countries. The World Bank, Washington D.C., p 76

    Google Scholar 

  • Wu H, Sparks C, Amoah B, Jones HD (2003) Factors influencing successful Agrobacterium-mediated genetic transformation of wheat. Plant Cell Rep 21:659–668

    PubMed  CAS  Google Scholar 

  • Yamamoto YT, Rajbhandari N, Lin X, Bergmann BA, Nishimura Y, Stomp AM (2001) Genetic transformation of duckweed Lemna gibba and Lemna minor. In Vitro Cell Dev Biol Plant 37:349–353

    Article  CAS  Google Scholar 

  • Yilmaz E, Akyurt I, Gunal G (2005) Use of duckweed, Lemna minor, as a protein feedstuff in practical diets for common carp, Cyprinus carpio. Fry Turk J Fish Aquat Sci 4:105–109

    Google Scholar 

Download references

Acknowledgements

GKC thanks to Council of Scientific and Industrial Research (CSIR), New Delhi, India for providing financial support. We are also thankful to Dr. Richard Jefferson, Center for Application of Molecular Biology to International Agriculture (CAMBIA), Australia, for providing the plasmid pCAMBIA2301.

Author information

Authors and Affiliations

  1. Centre for Biotechnology, Maharshi Dayanand University, Rohtak, 124001, India

    Gulshan Chhabra, Darshna Chaudhary, Manish Sainger & Pawan K. Jaiwal

Authors
  1. Gulshan Chhabra
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Darshna Chaudhary
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Manish Sainger
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Pawan K. Jaiwal
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Pawan K. Jaiwal.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Chhabra, G., Chaudhary, D., Sainger, M. et al. Genetic transformation of Indian isolate of Lemna minor mediated by Agrobacterium tumefaciens and recovery of transgenic plants. Physiol Mol Biol Plants 17, 129–136 (2011). https://doi.org/10.1007/s12298-011-0059-5

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12298-011-0059-5

Keywords

Search

Navigation