Unisexual Reproduction In Xenopus Tropicalis And Transcriptome Analysis Of Gynogenetic Haploids

Gynogenesis and androgenesis are two commonly used methods for artificial unisexual reproduction in vertebrates.Application of unisexual reproduction can greatly shorten the time for establishing gene modified animal lines.Gynogenesis is a kind of unisexual reproduction that requires the presence of sperm without the actual contribution of their DNA.The development of the embryo is controlled only by the maternal genes.Spontaneous gynogenesis in vertebrates is mostly found in some fish and a small number of amphibians.Artificially induced gynogenetic animals can be obtained through eliminating male inheritance by destroying DNA in sperm and restoring the chromosome diploidy by inhibiting the second meiosis or the first cleavage of eggs.Androgenesis refers to a special unisexual reproduction that relies solely on the male pronucleus to develop offsprings,while the eggs do not provide nuclear genetic material.Spontaneous androgenesis is extremely rare in vertebrates.Induced androgenetic animals in principle can be obtained by genetically inactivating the egg chromosomes and doubling the chromosomes of the sperm.In Xenopus tropicalis,artificial gynogenesis can be used to generate healthy and fertile gynogenetic diploid individuals,while there are still many unresolved problems to obtain healthy androgenetic frogs.In this study,artificial gynogenesis in X.tropicalis was stably established.Healthy diploid homozygotes were obtained and the survival rates of tadpoles from early cold shock gynogenesis and late cold shock gynogenesis were evaluated.In contrast,attempts to obtain androgenetic diploid individuals in X.tropicalis remained unsuccessful.Upon cold shock,the embryos all died before neurulation,indicating a sharp difference between gynogenesis and androgenesis in X.tropicalis.Although the haploid syndrome in amphibians has been noticed for a long time,the underlying molecular mechanisms are still inconclusive.In this study,RNA-Seq was used to identify differentially expressed genes(DEGs)between gynogenetic haploids and diploids.GO and KEGG analyses were performed on the DEGs.The data indicate that gynogenetic haploids display obvious abnormalities in glucose metabolism,transcriptional regulation of DNA,ubiquitination,and phosphorylation.The significant aberrant expression of MAPK signaling pathway and Fox O signaling pathway in haploids may trigger abnormal apoptosis.The abnormal expression of cytokine-cytokine receptor interaction,Toll-like receptor signaling pathway,C-type lectin receptor signaling pathway,etc.may lead to defects in the immune function of haploid embryos.Together,these data contribute to elucidating haploid syndrome in X.tropicalis.It has been reported that the deletion of p53 gene can improve the survival adaptability of several haploid cell lines,so that they can better maintain haploidy.However,the experimental results of my work showed that the deletion of p53 gene did not improve the viability at the embryo level,the mechanism of haploid syndrome in X.tropicalis needs to be further explored.