## 4 Discussion
### Stimulation of Celestial Goldfish Eggs by Distant Sperm  
There were few studies on the production of diploid female nuclear development offspring through distant hybridization. [PERSON] et al. [21] induced the development of diploid offspring by using heterologous sperm to prompt the development of scattered scale mirror carp female nuclei, with a success rate of 0.8548% for the appearance of diploid female nuclei development. [PERSON] et al. [22] induced the development of diploid female nuclei in large yellow croaker using non-inactivated light yellow croaker sperm, obtaining diploid female nuclei development and successfully producing normal and viable diploid female nuclei development offspring through pressure shock on the haploid fertilized eggs. However, in this experiment, using non-inactivated largemouth bass sperm to stimulate celestial goldfish eggs did not activate embryonic development, and the hybridization was unsuccessful. [PERSON] [23] summarized that offspring produced by intra-species hybridization were usually fertile, while inter-species hybridization was often difficult to succeed due to reproductive incompatibility. Largemouth bass (_Micropertus salmonides_), belonging to the order _Perciformes_, suborder _Percoidei_, family _Centrchidae_, genus _Micropertus_[24], and goldfish belonging to the order _Cypriniformes_, family _Cyprinidae_, genus _Carassius_[25], belong to different orders, and the hybridization might be unsuccessful due to their distant relationship and reproductive incompatibility.  
[PERSON] et al. [26] concluded that when the number of chromosomes in the parents was the same or the number of maternal chromosomes was greater than that of the paternal chromosomes, hybridization between them was compatible, and offspring would be obtained. For example, [PERSON] [27] conducted distant reciprocal crosses between goldfish (_Carassius auratus_, 2n = 100) and yellow-tail load (_Xenocyprisdavidi Bleeker_, 2n = 48). The hybridization of goldfish females with yellow-tail load males produced viable offspring with fertilization and hatching rates of 75.3% and 64.6%, respectively, while the reverse cross (goldfish males \(\times\) yellow-tail load females) did not produce viable offspring. Similar results were observed in the hybridization between grass carp [28], with the same chromosome number as Arctic char, both having 48 chromosomes and belonging to the same subfamily, and the hybridization between mandarin fish and largemouth bass, with 2n = 48 chromosomes, belonging to different families in the same order. In contrast, in this experiment, celestial goldfish, with 2n = 100 chromosomes [25], as the female parent and largemouth bass, with 2n = 46 chromosomes [29], as the male parent did not result in fertilization. After cold shock, the eggs did not develop, indicating that they were incompatible, possibly due to reproductive barriers. The above experiment result was inconsistent with the view of [PERSON] et al. [26], and the view of [PERSON] et al. [26] might be applicable to the same-order fish, while different-order fish might be incompatible due to their distant relationship and inability to fertilize and produce offspring.