Excision of a selectable marker gene in transgenic banana using a Cre/lox system controlled by an embryo specific promoter

Plant Mol Biol. 2013 Sep;83(1-2):143-52. doi: 10.1007/s11103-013-0058-8. Epub 2013 Apr 17.

Abstract

Antibiotic and herbicide resistance genes have been used in transgene technology as powerful selection tools. Nonetheless, once transgenic events have been obtained their presence is no longer needed and can even be undesirable. In this work, we have developed a system to excise the selectable marker and the cre recombinase genes from transgenic banana cv. 'Grande Naine' (Musa AAA). To achieve this, the embryo specific REG-2 promoter was isolated from rice and its expression pattern in banana cell clumps, somatic embryos and regenerated plantlets was characterized by using a pREG2::uidA fusion construct. Subsequently, the REG-2 promoter was placed upstream of the cre gene, conferring Cre functionality in somatic embryos and recombination of lox sites resulting in excision of the selectable marker and cre genes. PCR analysis revealed that 41.7 % of the analysed transgenic plants were completely marker free, results that were thereafter confirmed by Southern blot hybridization. These results demonstrate the feasibility of using developmentally controlled promoters to mediate marker excision in banana. This system does not require any extra handling compared to the conventional transformation procedure and might be useful in other species regenerating through somatic embryogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Genes, Plant*
  • Genetic Markers
  • Genetic Vectors / genetics
  • Genetic Vectors / metabolism
  • Globulins / genetics
  • Globulins / metabolism
  • Integrases / genetics
  • Integrases / metabolism*
  • Molecular Sequence Data
  • Musa / genetics*
  • Musa / metabolism
  • Oryza / genetics
  • Plant Proteins / genetics
  • Plant Proteins / metabolism
  • Plant Somatic Embryogenesis Techniques
  • Plants, Genetically Modified / genetics*
  • Plants, Genetically Modified / metabolism
  • Promoter Regions, Genetic*
  • Seeds / genetics*
  • Seeds / metabolism
  • Transformation, Genetic

Substances

  • Genetic Markers
  • Globulins
  • Plant Proteins
  • Cre recombinase
  • Integrases