Rivera Pagán, Ingrid T.

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    Chromosomal rearrangements of Psittacidae evolution: zoo-fish in Amazona vittata and in silico analysis of chromosomal rearrangements validated by de novo PCR
    (2014) Rivera Pagán, Ingrid T.; Martínez Cruzado, Juan C.; College of Arts and Sciences - Sciences; Oleksyk, Taras K.; Ortíz Acevedo, Alejandro; Department of Biology
    The Puerto Rican parrot (Amazona vittata, AVI) is a critically endangered species and the only native parrot species remaining on all U.S. territories. After its genome was sequenced through a community-funded project, we studied the ALLPATH 2012 genome assembly largest 3,099 scaffolds using UCSC Genome Browser’s BLAT tool against the chicken reference genome. The alignments of the first and last 25 kb of the scaffolds showed a total of 164 possible chromosomal rearrangements. A selection of the alignments with scores greater than 1,000 on both sides was made and primers were designed from the rearrangement joint regions. Out of the 19 putative rearrangements that did not involve sexual or unknown chromosomes, five were confirmed as true rearrangements in four Amazon parrots, including the Puerto Rican parrot. With our de novo PCR validation approach, we also confirmed that three of these rearrangements occurred before the evolutionary split of the African grey parrot, joining chicken chromosomes 2 with 15, 6 with 7, and 8 with 9. PCR further showed that the remainder two chromosomal rearrangements, joining chicken chromosomes 3 with 9 and 6 with 7, occurred after the split of the scarlet macaw. In order to revalidate these rearrangements, we used the well-established Fluorescent In Situ Hybridization (FISH) technique, hybridizing 10 chicken (GGA) whole-chromosome paints on Puerto Rican parrot metaphases. The FISH revealed three [6 with 7 (2) and 8 with 9] of the previously confirmed rearrangements through PCR and the rest of the chicken paints hybridized to unique segments or whole chromosomes in the parrot. Chicken chromosome probe 1 gave a signal on AVI3, GGA2 on AVI2 and a pair of microchromosomes, GGA3 on AVI1, GGA4 on AVI4 and to an arm of the metacentric AVI8, while GGA5 painted AVI7, GGA6 & 7 interestingly painted AVI6 in an alternated fashion, and GGA8 and 9 shared hybridization signals on a microchromosome pair. Furthermore, GGA9 also painted AVI9 and GGAZ painted its equivalent Z chromosome in the parrot. In addition, 100 of these metaphases were also used to describe A. vittata’s karyotype, which averaged a chromosome diploid number of 76. All together, these results contribute as the first complete description of the karyotype evolution of an Amazon species.