A method combining TA cloning and fluorescence screening for rapid acquisition of transgenic seeds

Biotechniques. 2020 May;68(5):251-256. doi: 10.2144/btn-2019-0141. Epub 2020 Mar 6.

Abstract

The establishment of transgenic plants has greatly promoted the progress of plant research. However, traditional selection methods using antibiotics or herbicides may miss any positive transformants with growth defects. Additionally, screening with antibiotics/herbicides requires a huge amount of seeds, sterile work conditions and a large amount of space to germinate plants, making the selection process time- and labor-consuming. In this study, we constructed a novel stable transformation vector, plasmid of OLE1-GFP T-DNA vector (pOGT), which can shorten the steps of cloning foreign genes into expression vectors by using TA cloning. Additionally, selection of transformed seeds with fluorescence overcomes the difficulties of conventional selection with antibiotics/herbicides and simplifies the screening process for transgenic plants.

Keywords: Arabidopsis thaliana; OLE1 gene; TA cloning; green fluorescent protein (GFP); selection marker; stable expression vector.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Arabidopsis / classification
  • Arabidopsis / genetics
  • Cloning, Molecular / methods*
  • Genes, Plant / genetics*
  • Genetic Vectors / genetics
  • Membrane Proteins / genetics
  • Plant Proteins / genetics
  • Plants, Genetically Modified* / classification
  • Plants, Genetically Modified* / genetics
  • Plasmids / genetics
  • Seeds* / classification
  • Seeds* / genetics

Substances

  • Membrane Proteins
  • Plant Proteins