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The Hefang crucian carp (HFJ) derived from Carassius cuvieri (♀) × C. auratus red var. (♂) and the crucian carp-like homodiploid fish (NCRC) derived from Cyprinus carpio (♀) × Megalobrama amblycephala (♂) are important germplasm resources of crucian carp. To investigate the population characteristics and reveal the current genetic situations of these hybrid varieties, we sequenced one nuclear molecular marker transferrin (Tf) among six Carassius populations, which included two hybrid varieties (HFJ and NCRC), one laboratory variety (red crucian carp, RCC) and three wild populations. A total of 166 distinct Tf alleles (A1∼A166) were identified, with sequence lengths ranging from 1145 bp to 1244 bp. Unexpectedly, the nucleotide diversity of Tf alleles in cultured populations (HFJ, NCRC, and RCC) exceeded that of wild populations. The phylogenetic reconstruction analysis results showed that HFJ and C. cuvieri were closely related, but they could not be separated from C. auratus. Besides, while most Tf alleles of NCRC and other Carassius populations were intermixed in five clades, two Tf alleles, together with Cy. carpio, formed a distinct monophyletic clade. Moreover, network and genetic structure analyses failed to distinguish between wild and cultured populations. Although AMOVA analysis based on Tf alleles indicated genetic differentiation among all six populations, the majority of the genetic variation (92.00 %) was observed within populations. Furthermore, the population dynamics analysis suggested that these populations have maintained relative stability in recent history. Therefore, the elevated nucleotide diversity of Tf alleles in cultured populations, the ambiguous population structure, and the limited genetic divergence among the six Carassius populations highlighted the hybrid genetic characteristics of HFJ and NCRC. These findings, based on Tf alleles, differ from the results obtained using mitochondrial gene markers, providing an alternative perspective for germplasm evaluation of hybrid varieties and crucian carp breeding.

Cryopreservation of sperm is an effective method for conserving germplasm resources in fish genetic breeding. The high-quality hybrid fish (WR) derived from white crucian carp (Carassius cuvieri, WCC, ♀) and red crucian carp (C. auratus red var., RCC, ♂), possesses valuable traits such as high survival rates, strong resistance, and rapid growth, representing an important germplasm resources of crucian carp. This study compared the effects of different antifreeze solutions on sperm viability among three varieties (WR, WCC, and RCC) and examined changes in enzyme activity, fertilization rates, and hatching rates after cryopreservation, aiming to enhance the cryogenic sperm cryopreservation technique in hybrid fish and investigate the mechanisms underlying spermatozoa damage caused by cryopreservation. The results showed that the antifreeze combination of D14 with 15 % dimethyl sulfoxide (DMSO) had the best effect in preserving the sperm of WR and WCC, while D20 with 10 % DMSO was the optimal combination for RCC sperm. After ultra-low temperature preservation, the longevity, fertilization, and hatching rates of frozen sperm were significantly lower (P

The surprising variation in the number of Hox clusters and genome structure in ray-finned fish lineages reflects the history of duplications and subsequent lineage-specific gene loss. However, there are few studies on whether Hox clusters in the early generations of hybrid lineages show more significant variation due to the continuous genomic oscillation caused by distant hybridization. We sequenced and analyzed Hox gene clusters from a crucian carp-like homodiploid fish (NCRC) lineage (a new hybrid lineage derived from common carp (Cyprinus carpio) (♀) ​× ​blunt snout bream (Megalobrama amblycephala) (♂)). In the NCRC lineage, we reconstructed seven Hox clusters consisting of 48 Hox genes, ten of which were pseudogenes. The number of putative Hox clusters generated in NCRC-F1 was increased greatly by distant hybridization to an average number almost twice that in the maternal parent. This increasing trend continued in the subsequent self-mating generations of NCRC-F1. In contrast, the number of Hox cluster fragments inherited from the original parents gradually decreased as the number of NCRC lineage generations increased. This pattern was also found in the inheritance of recombinant Hox clusters. In terms of base composition, some genetic rules for the inheritance of these Hox clusters between different generations of the NCRC lineage were identified. Furthermore, the newly derived mutated Hox clusters in the NCRC lineage showed phylogenetic relationships that were closer to either crucian carp or silver crucian carp, revealing a clear evolutionary path. This study deepens our understanding of the evolution of Hox genes in the ray-finned fish clade.