fth1b基因敲除对斑马鱼咽齿早期矿化的影响研究

Knockout fth1b affects early mineralization of zebrafish pharyngeal teeth

目的探索铁蛋白重肽基因(fth1b)在斑马鱼咽齿发育过程中的表达模式及其功能,为后续牙齿相关研究奠定基础。方法收集受精后56、72、96及120 h斑马鱼胚胎,运用整胚原位杂交技术检测fth1b基因在斑马鱼咽齿发育过程中的表达;并运用规律成簇间隔短回文重复序列(CRISPR)/Cas9基因编辑技术对fth1b基因特异性敲除,在fth1b-/-突变体中检测斑马鱼咽齿的发育状况。结果fth1b基因与已知斑马鱼咽齿标记基因dlx2b表达模式类似,特异性表达于斑马鱼咽齿发育过程中;靶向性敲除fth1b基因后,早期发育过程中,斑马鱼咽齿标记基因pitx2、dlx2b表达与野生型无较大差别,而咽齿的矿化较野生型斑马鱼弱。结论fth1b基因特异性表达于斑马鱼咽齿,对斑马鱼咽齿的早期矿化起作用。

Objective A study was conducted to explore the expression pattern and function of ferritin heavy polypeptide gene(fth1b)in zebrafish pharyngeal teeth development and lay the foundation for subsequent research on teeth development and mineralization.Methods The zebrafish embryos were harvested at 56,72,96,and 120 h after fertilization.The expression of fth1b in zebrafish pharyngeal teeth development was detected by whole embryo in situ hybridization and compared with the known pharyngeal teeth marker dlx2b.The specific knockout of fth1b gene was performed using clustered regularly interspaced short palindromic repeats(CRISPR)/Cas9 gene editing technology.The development of zebrafish pharyngeal teeth was detected in the fth1b^-/- mutant.Results The expression pattern of fth1b gene was very similar to that of the known zebrafish pharyngeal teeth marker dlx2b and was specifically expressed in the zebrafish pharyngeal teeth during development.After the specific knockout of the gene fth1b,the earliest gene that can be detect in zebrafish pharyngeal teeth-pitx2 was expressed normally during early development.The dlx2b expression was not significantly different from that of wild type zebrafish,but the mineralization of pharyngeal teeth in the mutant was weaker than that of wild type zebrafish.Conclusion The gene fth1b is specifically expressed in zebrafish pharyngeal teeth and acts on their early mineralization.