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High Frequency of T-DNA Deletions in Transgenic Plants Transformed with Intron-Containing Hairpin RNA Genes

Abstract

The transcriptional activator protein gene, the AC4 gene and the common region sequences of Mungbean yellow mosaic virus were cloned in hairpin RNA (hpRNA) cassettes in Agrobacterium binary vectors. Of the 33 transgenic tobacco plants generated by transformation with five different binary plasmids with three hpRNA constructs, only 12 transgenic plants had the intact T-DNAs which comprised the selectable marker gene (SMG) and the complete hpRNA gene. Fourteen plants harboured the SMG part of the T-DNA but lacked the hpRNA part. Seven plants had the SMG but had only truncated hpRNA genes. Ten of the 12 plants with the complete hpRNA genes also had many additional truncated T-DNA copies. Southern blot analysis of the AC4 hpRNA-transformed plants with the bar and AC4 probes, which flanked left and right T-DNA borders, respectively, revealed that the bar gene portion was intact in seven out of eight plants, whereas the AC4 hpRNA portion was truncated in seven out of eight plants. Our findings revealed that the T-DNAs with hpRNA genes were highly prone for deletions in transgenic plants.

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Acknowledgements

The authors thank Dr. Thomas Hohn (University of Basel, Switzerland) for the hpCR and hpTrAP cassettes. Ms. Bridget Jeyatha is acknowledged for the construction of pGA-hpCR. We acknowledge the research support from the Department of Biotechnology, Government of India and University Grants Commission, Government of India. S. Sunitha and P. V. Shivaprasad acknowledge the Council for Scientific and Industrial Research, Government of India for their research fellowships.

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Correspondence to Karuppannan Veluthambi.

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Sunitha, S., Shivaprasad, P.V., Sujata, K. et al. High Frequency of T-DNA Deletions in Transgenic Plants Transformed with Intron-Containing Hairpin RNA Genes. Plant Mol Biol Rep 30, 158–167 (2012). https://doi.org/10.1007/s11105-011-0327-0

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Keywords

  • hpRNA genes
  • MYMV-AC4
  • MYMV-CR
  • MYMV-TrAP
  • T-DNA truncation