3 research outputs found
A protocol for Agrobacterium-mediated transformation of Kalanchoë blossfeldiana with a flavonoid 3',5' hydroxylase (F3'5'H) gene
In the present investigation, explants from Kalanchoë blossfeldiana were used for gene transformation. The young leaves were inoculated with Agrobacterium tumefaciens LBA4404 strain with a binary vector plasmid pArtblue containing F3'5'H gene under control of CaMV35S promoter and nptII selectable marker gene. After inoculation, the explants were transferred to the co-cultivation medium. They were then transferred to the selection medium containing kanamycin and were sub-cultured every two weeks. Leaves of the putative transgenic shoots that survived in the selection medium were used in reverse transcription polymerase chain reaction (RT-PCR) analysis to detect gene expression. The RT-PCR analysis showed the presence of 550 bp F3'5'H amplification products and had an expression of F3'5'H gene. Plants with the introduced F3'5'H gene produced totally pale red flowers.Key words: Kalanchoë blossfeldiana, Agrobacterium-mediated transformation, young leaf, F3'5'H gene, reverse transcription-polymerase chain reaction (RT-PCR)