Banana (Musa sp.) is considered as one of the most important fruit crops worldwide
as well as in Egypt. The main goal of this study was to construct the open reading frame (ORF)
of banana bunchy top nanovirus (BBTV)-DNA-3 that encodes the viral coat protein (cp) gene
for banana transformation. The previously sequenced BBTV-G-DNA-3-ORF that cloned into
plasmid pH1 was used as a template for PCR amplification using two specific primers containing
Bam H1 site. A new plasmid called pRHA1 containing the amplified ORF under the control
of maize polyubiquitin (ubi) promoter was created. The bar gene (herbicide-resistance gene as a selectable marker) cassette (bar gene, Cauliflower mosaic caulimovirus (CaMV) 35S promoter
and nos terminator) was released from plasmid pAB6 using Hind III-digestion and subcloned
into the Hind III-digested plasmid pRHA1 to create the plasmid pRHA2 via the microprojectile
bombardment transformation system. The plasmid pRHA2 was successfully introduced in the
applied banana cultivar. Leaf painting test was conducted to confirm the expression of the
bar gene in the putative transformed banana lines. The presence and expression of BBTV-Gcp
gene were also detected using some molecular (polymerase chain reaction and dot blot using
a cold DNA probe) and serological (ELISA and western blot) techniques, respectively, in the
obtained transgenic banana lines.