Bromoxynil resistant tobacco (Nicotiana tabacum L) generation using a synthetic bxn gene

Synthetic biology is developing fast and resulting in the preparation of various in vitro made DNA sequences with modified specifications. Transgenic tobacco plants resistant to the nitrile class bromoxynil herbicide were produced applying a synthetically prepared bxn gene. Bromoxynil’s effects on postemerged tobacco plantlets were assessed before genetic transformation, on which leaf necrosis and rigorous damage were observed. Genetic transformation of tobacco was carried out using regenerated shoot explants from 10-day-old cotyledon cultured in MS medium. Regenerated shoot explants were inoculated and cocultured with Rhizobium radiobacter strain LBA 4404 containing recombinant pCAMBIA (bxn+). Hygromycin resistant shoot explants were developed and transferred into selective rooting medium, followed by soil acclimatization. The existence and integration of the synthetic bxn gene in the transgenic tobacco genome were examined and proved by PCR and nonradioactive Southern blotting techniques. PCR analysis of transformants revealed the expected size of 1063 bp synthetic bxn gene in the transgenic tobacco genome. Southern blotting showed a clear and explicit image of an integrated bxn gene band in the transformed plant genome. The herbicide resistant bioassay test showed the resistance of transformants for a minimum concentration of 50 mg of bromoxynil dissolved in 200 mL of water. This experiment suggested that the transgenic tobacco plantlets are resistant to the herbicide bromoxynil.