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Organ regenerative ability depends on the animal species and the developmental stage. The molecular bases for variable organ regenerative ability, however, remain unknown. Previous studies have identified genes preferentially expressed in the blastema tissues in various animals, but transcriptome analysis of the isolated proliferating blastema cells has not yet been reported. In the present study, we used RNA-sequencing analysis to analyze the gene expression profile of isolated proliferating blastema cells of regenerating  Xenopus laevis  tadpole tails. We used flow cytometry to isolate proliferating cells, and non-proliferating blastema cells, from regenerating tadpole tails as well as proliferating tail bud cells from tail bud embryos, the latter two of which were used as control cells, based on their DNA content. Among the 28 candidate genes identified by RNA-sequencing analysis, quantitative reverse transcription-polymerase chain reaction identified 10 genes whose expression was enriched in regenerating tadpole tails compared with non-regenerating tadpole tails or tails from the tail bud embryos. Among them, whole mount  in situ  hybridization revealed that  chromosome segregation 1-like  and  interleukin 11  were expressed in the broad area of the tail blastema, while  brevican ,  lysyl oxidase , and  keratin 18  were mainly expressed in the notochord bud in regenerating tails. We further combined whole mount  in situ  hybridization with immunohistochemistry for the incorporated 5-bromo-2-deoxyuridine to confirm that  keratin 18  and  interleukin 11  were expressed in the proliferating tail blastema cells. Based on the proposed functions of their homologs in other animal species, these genes might have roles in the extracellular matrix formation in the notochord bud ( brevican  and  lysyl oxidase ), cell proliferation ( chromosome segregation 1-like  and  keratin 18 ), and in the maintenance of the differentiation ability of proliferating blastema cells ( interleukin 11 ) in regenerating tadpole tails.

Unique Gene Expression Profile of the Proliferating Xenopus Tadpole Tail Blastema Cells Deciphered by RNA-Sequencing Analysis