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In our first study, we have analyzed more than 2,500 nucleotide sequences from the ITS region of 87 accessions representing the family Musaceae, including 54 Musa diploids representing all sections of the Musa genus including four inter-subspecific hybrids, two Ensete and one Musella species. In addition to the representatives of wild Musa species, ITS region of 30 hybrid banana clones was analyzed. Three approaches were used to obtain ITS sequences. As the first step, the ITS region of all diploid genotypes was amplified using specific primers and four PCR products from individual PCR reactions of each accession were used for direct Sanger sequencing. Only 23 diploid genotypes produced readable ITS sequences with no polymorphism, the remaining diploid accessions produced polymorphic ITS sequences. The ITS region of heterogeneous diploids and all hybrid clones was cloned and 15 - 85 DNA clones bearing ITS region were sequenced. Finally, we have assessed the usefulness of massively parallel 454 sequencing for the analysis of the ITS loci.
The ITS sequence region is now beeing also used for detailed charactreization of phylogeentic relationships of other, wild diploid species as well as Musa diploid and triploid cultivars.
