3.5 Chromosomal breakage and reunion verified by synteny
comparison
To verify the hypothesis of chromosome breakage and reunion, comparative
chromosome analysis was performed using MUMmer4 to identify genome-scale
syntenic regions between female and male O. bidens . The genome
sequences corresponding to their chromosomes were highly consistent
(Figure 6). Similarly, the chromosomal sequences of female GH14/GH38
exhibited a significant identity with the male LG06 using MUMmer4 with a
minimum match length of 1,000 bp and 85% of filter (Figure 7a). The
female GH14/GH38 chromosomal sequence alignment to the male LG06 showed
identities of 97.93% and 97.72%, respectively, suggesting that the
female GH14 and GH38 have fused to form the male LG06. Indeed,
structural variations were identified by performing sequence
comparisons, and we found that in the male LG06, the deletions included
232 in the intergenic region, 20 in the upstream region, 12 in the
downstream area, and 259 in the intronic region. The insertion sites
included 218 in the intergenic region, 26 in the upstream region, 23 in
the downstream region, and 248 in the intronic area. Finally, the
duplication, inversion, and translocation sites lacked a homolog to the
female GH14/GH38 (Figure 7b). The deletions and insertions that occurred
in the male LG06 were lower. They accounted for 3.4% (1038/30202) of
the entire variable sites in all-maleO.
bidens chromosomes, suggesting that the male LG06 resulted from the
fusion of the female GH14 and GH38 rather than from massive
rearrangements.
According to the annotated gene sets in the male O. bidensgenome, using homology searching of the female O. bidens genome,
we identified seven male-specific genes in the male LG06 chromosome,
including four uncharacterized protein genes; two genes involved in
replication, recombination, and repair functions (EVM0022038,
EVM0026331); and one gene annotated as DNA polymerase epsilon subunit 2
(EVM0017949), engaged in DNA-dependent DNA replication in biological
processes.
To reveal the possible mechanism of chromosome number variation by
breakage in Cyprinid fish, the chromosomes of male O. bidens were
aligned to a single chromosome of C. idella , exhibiting
significant inter-chromosomal conservation (Figure 8a). C. idellashowed a high homology and 269 synteny blocks with 21,628 gene pairs in
the genome comparison. Each of the 14 C. idella chromosomes broke
into two chromosomes of the male O. bidens , and massive
inter-chromosomal rearrangements occurred after the divergence of the
two species (Figure 8b). Notably, the fusion of the broken fragments
from C. idella Ch08 and Ch17 chromosomes gave rise to the LG06
chromosome of male O. bidens , indicating that chromosome
breakages help to increase their number in fish chromosomal evolution.