新生隐球菌STE12α基因的克隆及表达载体的构建

Cloning and construction expression plasmid of STE12α gene in Cryptococcus neoformans

目的从新生隐球菌的基因组中扩增出STE12α基因,并构建相应的表达载体,以进一步研究STE12α基因对隐球菌的生长特性及致病性的影响。方法采用PCR方法以及基因重组方法扩增并克隆新生隐球菌基因组中的STE12α基因,建立具有表达野生型STE12α基因的表达载体。结果从新生隐球菌的基因组获得STE12α全基因,建立重组子pUCm-STE12α/NovaBlue以及重组表达载体质粒pGAPZ-STE12α,实现了STE12α基因的转化并获得表达。结论成功地克隆了新生隐球菌STE12α基因并构建了可表达野生型STE12α基因的表达载体,为进一步研究STE12α基因功能打下了良好的基础。

Objective To clone and construct expression plasmid of STE12α gene in Cryptococcus neoformans. Methods PCR technique and recombination metheds were used in cloning and construction expressing plasmid of STE12α gene. Results STE12α gene was amplificated from genomic DNA of Cryptococcus neoformans strain and cloned into pUCm-T vector. EcoRI-NotI fragment of pUCm-STE12α containing STE12α gene coding region was inserted into the same EcoRI-Notl site of pGAPZalpha-A vector to yield the pGAPZ-STE12α expressing plasmid. RT-PCR showed that STE12α gene was expressed in Cryptococcus neoformans transformants. Conclusion Successful cloning STE12α gene and construction wild STE12α gene expression plasmid will be useful for further pathogenic research of Cryptococcus neoformans .