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<p>This study aimed to develop a reliable and efficient genetic transformation method for the ornamental Indian Lotus (<i>Nelumbo nucifera</i> Gaertn.) using the sonication-assisted <i>Rhizobium radiobacter-</i>mediated transformation technique. To conduct the transformation, shoot apical meristem explants were infected with <i>Rhizobium radiobacter</i> (synonym <i>Agrobacterium tumefaciens)</i> strain LBA 4404 containing a binary vector pBI121 that harbours the GUS reporter gene (<i>uid</i>A) and kanamycin resistance gene <i>npt</i>II for plant selection. To improve the transformation efficiency, we optimized parameters such as bacterial cell density, sonication duration, infection time, co-cultivation duration, acetosyringone concentration, cefotaxime, and kanamycin concentrations. Sonication treatment at 42&#xa0;kHz for 90&#xa0;s recorded the highest transformation efficiency. The selection of regenerated plantlets was performed on a kanamycin-supplemented selection medium. The putative transformants showed GUS expression in the leaves and petioles. The presence of the GUS gene was also confirmed in the putative transformants through PCR, with the appearance of the expected amplicon size of 520&#xa0;bp. The presence of <i>npt</i>II was confirmed by PCR in the putatively transformed plants with an amplicon size of 530&#xa0;bp. The maximum regeneration frequency obtained was 72.66%, and the highest transformation efficiency achieved was 9.0% in the Indian Lotus.</p> Graphical abstract <p></p>

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Sonication-assisted Rhizobium radiobacter-mediated genetic transformation of Indian Lotus (Nelumbo nucifera Gaertn.)

  • 期刊论文

  • 发表日期：

    2025年1月7日

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  • Rita Verma,
  • Anshu Sahu,
  • Rajan Kumar Gupta,
  • Indraneel Sanyal

摘要

This study aimed to develop a reliable and efficient genetic transformation method for the ornamental Indian Lotus (Nelumbo nucifera Gaertn.) using the sonication-assisted Rhizobium radiobacter-mediated transformation technique. To conduct the transformation, shoot apical meristem explants were infected with Rhizobium radiobacter (synonym Agrobacterium tumefaciens) strain LBA 4404 containing a binary vector pBI121 that harbours the GUS reporter gene (uidA) and kanamycin resistance gene nptII for plant selection. To improve the transformation efficiency, we optimized parameters such as bacterial cell density, sonication duration, infection time, co-cultivation duration, acetosyringone concentration, cefotaxime, and kanamycin concentrations. Sonication treatment at 42 kHz for 90 s recorded the highest transformation efficiency. The selection of regenerated plantlets was performed on a kanamycin-supplemented selection medium. The putative transformants showed GUS expression in the leaves and petioles. The presence of the GUS gene was also confirmed in the putative transformants through PCR, with the appearance of the expected amplicon size of 520 bp. The presence of nptII was confirmed by PCR in the putatively transformed plants with an amplicon size of 530 bp. The maximum regeneration frequency obtained was 72.66%, and the highest transformation efficiency achieved was 9.0% in the Indian Lotus.

Graphical abstract