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Agrobacterium–mediated genetic transformation of Elymus breviaristatus with Pseudomonas pseudoalcaligenes insecticidal protein gene

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Abstract

An efficient and repeatable protocol for generating transgenic Elymus breviaristatus plants was first established by Agrobacterium-mediated transformation: calli of Elymus breviaristatus were induced from mature seeds, and proliferated on callus induction and maintenance medium containing kinetin 0.05 mg/l and 2, 4-D 8.0 mg/l under dim-light condition (10–20 μmol/m2s,16 h light) at 26°C. The calli produced were used for transformation mediated by Agrobacterium tumefaciens, EHA105, which carries the binary vector pCAMBIA 1304-ppip, coding for the Pseudomonas pseudoalcaligenes insecticidal protein gene (ppip) and hygromycin phosphotransferase II gene (hptII). The calli transformed were selected and grown in the presence of 60 mg/l hygromycin. Approximately 90 transgenic plants were produced, and transformation frequency of the calli reached 10.52%. The presence of ppip gene in the genomic DNA of regenerated plants was detected by means of PCR and Southern-blot analysis, and the expression of the transgenes was verified by reverse transcription-PCR. Approximately 90% of the tested plants appeared to have only one or two copies of the T-DNA inserts. Verification of anti-grasshoppers activity shows the mortality of grasshoppers reached 16.73%. These results indicated that Agrobacterium-mediated transformation in our condition was effective for transferring foreign genes into Elymus breviaristatus.

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Abbreviations

ABA:

abscisic acid

AS:

acetosyringone

6-BA:

6-benzylaminopurine

2,4-D:

2,4-dichlorophenoxyacetic acid

hptII:

hygromycin phosphotransferase II

MS:

Murashige and Skoog medium

NAA:

1-naphthaleneacetic acid

ppip :

seudoalcaligenes insecticidal protein gene

References

  • Abe T, Futsuhara Y (1985) Efficient plant regeneration by somatic embryogenesis from root callus of rice (Oryza sativa L.). J Plant Physiol 121:111–118

    CAS  Google Scholar 

  • Akutsu M, Ishizaki T, Sato H (2004) Transformation of the monocotyledonous Alstroemeria by Agrobacterium tumefaciens. Plant Cell Rep 22(8):561–568

    Article  PubMed  CAS  Google Scholar 

  • Batra S, Kumar S (2003) Agrobacterium-mediated transient GUS gene expression in buffel grass (Cenchrus ciliaris L.). J Appl Genet 44(4):449–458

    PubMed  Google Scholar 

  • Bettany AJ, Dalton SJ, Timms E, Manderyck B, Dhanoa MS, Morris P (2003) Agrobacterium tumefaciens-mediated transformation of Festuca arundinacea (Schreb.) and Lolium multiflorum (Lam.). Plant Cell Rep 21(5):437–444

    PubMed  CAS  Google Scholar 

  • Chai BF, Liang AH, Wang W, Hu W (2003) Agrobacterium-mediated Transformation of Kentucky Bluegrass. Acta Botanica Sinica 45(8):966–973

    Google Scholar 

  • Chai ML, Wang BL, Kim JY, Lee JM, Kim DH (2003) 2003 Agrobacterium-mediated transformation of herbicide resistance in creeping bentgrass and colonial bentgrass. J Zhejiang Univ Sci 4(3):346–351

    Article  PubMed  CAS  Google Scholar 

  • Chaudhry B, Yasmeen A, Husnain T (1999) Mini-scale genomic DNA extraction from cotton. Plant Mol Biol Rep 17:1–7

    Article  Google Scholar 

  • De Cleene M, Deley J (1976) The host range of crown gall. Bot Rev 42:389–466

    Article  Google Scholar 

  • Delannay XBJ, LaVallee RK, Proksch RL, Fuchs SR (1989) A Fischoff Field performance of transgenic tomato plants expressing the Bacillus thuringiensisvar. kurstaki insect control protein. Biotechnology 7:1265–1269

    Google Scholar 

  • Ding Shihua, Yang Zhirong (2001) N-Terminal analysis and antibody preparation of insecticidal protein from Pseudomonas pseudoalcaligenes. Acta Microbiologica Sinica 41(5):642–645

    Google Scholar 

  • Koziel MG, Beland GL, Bowman C, Carozzi NB, Crenshaw R (1993) Field performance of elite transgenic maize plants expressing an insecticidal protein derived from Bacillus thuringiensis. Bio/Technology 11:194–200

    Article  CAS  Google Scholar 

  • Li DX, Zhang J, Zhao J, Zhang Y, L1 L, Liu SJ, Chen F, Yang ZR (2006) Transformation of calli of siberian wildrye grass (Elymus sibiricus L. cv. chuancao No.2) mediated by Agrobacterium. J Plant Physiol Mol Biol 32(1): 45–51

    Google Scholar 

  • Liu SG, Zhu W, Yang ZR, Ge SR, He XY (1995) Isolation and identification of a pathogen of grasshoffers. Acta Microbiologica Sinica 35(2):86–90

    PubMed  CAS  Google Scholar 

  • McDonnell RE, Conger BV (1984) Callus induction and plantlet formation from mature embryo explants of Kentucky bluegrass. Crop Sci 24:573–578

    Article  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  • Nandakumar R, Chen L, Rogers SM (2005) Agrobacterium-mediated transformation of the wetland monocot Typha latifolia L. (Broadleaf cattail). Plant Cell Rep 23(10–11):744–750

    Article  PubMed  CAS  Google Scholar 

  • Rahman SML, Ebina M, Nakagawa H (2000) Agrobacterium tumefaciens-mediated gene transformation of Japanese lawn grass (Zoysia japonica Steud.). Plant & Animal Genome VIII Program January (9–12) San Diago, CA, USA (http://www.intl-pag.org/pag/8/ abstracts /pag8824.html.)

  • Wang SY, Yun JF, Jiang SH, Ha SQMG (2003) Inflorescence culture of interspecific hybrid F1 of Elymus canadensis × E. sibiricus. Grassland of China 25:61–64

    Google Scholar 

  • Wang ZY, Ge Y (2005) Agrobacterium-mediated high efficiency transformation of tall fescue (Festuca arundinacea). J Plant Physiol 162(1):103–113

    Article  PubMed  CAS  Google Scholar 

  • Wunn J, Kloti A, Burkhardt PK (1996) Transgenic indica rice breeding line IR58 expressing a synthetic crylA (b) gene from Bacillus thuringiensis provides effective insect pest control. Biotechnology 14:171–176

    Article  PubMed  CAS  Google Scholar 

  • Vasil IK (1988) Progress in the regeneration and genetic manipulation of cereal crops. BioTechnology 6:397–402

    Article  Google Scholar 

  • Vishnu Bhat, Sue J. Dalton, Suresh Kumar, Basrur Venkatesh BHAT, Madan G. Gupta, Phillip Morris (2001) Particle-inflow-gun-mediated genetic transformation of buffel grass (Cenchrus ciliaris L.): optimizing biological and physical parameters. J Applied Genetics 42(4):405–412

  • Yang H, Li JX, Guo SD (1989) Transgenic rice plants produced by direct uptake of δ-endotoxin protein gene from Bacillus thuringiensis into rice protoplasts. Sci Agric Sin 22(6):1–5

    CAS  Google Scholar 

  • Zhu ZQ, Wang JJ, Sun CS, Xu ZH, Zhu ZY, Yin GC, Bi FY (1975) Establishment of an efficient medium for anther culture of rice through comparative experiment on the nitrogen sources. Sci Sin 18:659–668

    Google Scholar 

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Acknowledgement

This work was supported by project (030451012-JY03-B31) from the Key Technologies R & D Programme of China.

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Authors and Affiliations

  1. Sichuan Academy of Grassland Science, 611731, Chengdu, P.R. China

    Daxu Li, Shiqie Bai, Minghong You, Gang Liu, Yu Zhang & Changbing Zhang

  2. College of Life Sciences, Key Laboratory of Bio-resources and Eco-environment, The Ministry of Education, Sichuan University, 610064, Chengdu , P.R. China

    Qun Sun, Min Huang, Jie Zhang, Linyong Zheng, Jian Zhao, Dunlian Qiu, Li Li, Zhirong Yang & Shaocai Li

Authors
  1. Daxu Li
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  2. Qun Sun
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  3. Min Huang
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  4. Jie Zhang
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  5. Shiqie Bai
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  6. Linyong Zheng
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  7. Jian Zhao
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  8. Dunlian Qiu
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  9. Li Li
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  10. Zhirong Yang
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  11. Minghong You
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  12. Gang Liu
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  13. Yu Zhang
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  14. Changbing Zhang
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  15. Shaocai Li
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Correspondence to Shaocai Li.

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Li, D., Sun, Q., Huang, M. et al. Agrobacterium–mediated genetic transformation of Elymus breviaristatus with Pseudomonas pseudoalcaligenes insecticidal protein gene. Plant Cell Tiss Organ Cult 89, 159–168 (2007). https://doi.org/10.1007/s11240-007-9229-0

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  • DOI: https://doi.org/10.1007/s11240-007-9229-0

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