应用改良的原位杂交方法分析Pax3基因在鸡胚脊髓中的表达

目的改良传统的原位杂交方法,提高其信号强度和特异性;应用改良的原位杂交方法分析Pax3基因在鸡胚脊髓中的表达情况。方法分子克隆Pax3 cDNA进入pCR-TopoII质粒,使用sp6 RNA聚合酶体外合成Pax3 cRNA反义探针,组织原位杂交分析Pax3(pairedbox3)mRNA在鸡胚脊髓内分布。结果获得高质量的Pax3 cRNA探针,通过改进的原位杂交方法发现Pax3 mRNA在鸡胚脊髓内呈区域状特异分布。结论改进的原位杂交方法显著提高了杂交信号特异性,Pax3在鸡胚脊髓内区域状表达显示Pax3基因在脊髓发育过程中扮演一定角色。

Expression of Pax3 gene in Chick Embryo spinal cord analyzed with the improved in situ hybridization

Objective To improve the traditionary procedure of in situ hybridization in order to enhance signal intensity and speciality,then to analyze the expression of Pax3(pairedbox3) in Chick Embryo spinal cord with the improved method.Methods Molecular cloning of Pax3 into pCR-TopoII,Pax3 cRNA antisense probes synthesis with sp6 RNA polymerase in vitro,and the distribution of Pax3 mRNA was analyzed by the improved in situ hybridization.Results Pax3 cRNA probe was obtained.Pax3 mRNA was found that it appeared as region specifically according to the improved method.Conclusion The improved method enhance remarkably the signal speciality of in situ hybridization.The expression of Pax3 mRNA appeares as region specifically in Chick Embryo spinal cord,which indicates that Pax3 plays certain role during the development of spinal cord.