Identification of 45S rDNA in Passiflora using low coverage sequencing: analysis of GC content and chromosomal localization.

Background: The 45S rDNA is considered the most useful chromosomal marker for cytogenetic analysis of Passiflora. Amplification of 45S rDNA sequence via PCR are more advantageous than sequence maintenance in vectors for chromosomal hybridization via FISH. We aimed both to identify 45S rDNA by sequencing data for chromosomal localization and to verify the relationship between GC content and CMA/DAPI banding.

Methods And Results: Low-coverage sequencing of Passiflora alata, P. cincinnata, and P. edulis was performed, and 45S rDNA units were identified using RepeatExplorer. The 45S rDNA units were used to construct a neighbor-joining tree to verify the similarities between the three species' 18S and 26S rDNA sequences. Clusters (CL)116 (P. alata), CL71 (P. cincinnata), and CL116 (P. edulis) were remarkably similar among the three species, and the 26S rDNA sequences of the clusters were similar to those of Populus tremuloides, Salix interior, and Averrhoa carambola (98% identity). The 26S rDNA was cytologically localized in the chromosomes of P. edulis, P. bahiensis, and the backcrossed hybrid (P. sublanceolata vs. HD13). The hybridization transfer capacity was evaluated in Citrus sunki and Cucumis melo. Finally, a chromosomal pair with a heteromorphic 26S rDNA site was observed in P. edulis, which was the same to that observed for CMA.

Conclusion: The amplification of the 26S rDNA in Passiflora via PCR and the chromosomal localization in Passiflora and other plant species was successfully achieved. The CMA bands were found to be related not only to the amount of GC but also to its structure and the number of repetitions.