灰黄青霉Pa-pex11基因对青霉素产生的影响  

Effects of Pa-pex11 gene on penicillin production in Penicillium aurantiogriseum

作　　者：杨晶[1,2] 安洋[1,2] 徐欣欣[1,2] 刘钢[1] 

机构地区：[1]中国科学院微生物研究所中国科学院真菌地衣系统学实验室,北京100101 [2]中国科学院研究生院,北京100049

出　　处：《微生物学报》2009年第11期1477-1482,共6页Acta Microbiologica Sinica

基　　金：国家科技支撑计划(2007BAI26B01)~~

摘　　要：【目的】研究灰黄青霉Pa-pex11对青霉素产生的影响。【方法】通过保守序列设计引物从灰黄青霉中克隆了含有pex11同源基因DNA片段,进而通过热不对称交错PCR(Thermal asymmetric interlacedPCR,TailPCR)扩增得到灰黄青霉中Pa-pex11基因的全序列。利用根癌农杆菌介导的遗传转化(ATMT)系统,在灰黄青霉基因组上导入额外的Pa-pex11基因。对灰黄青霉野生型菌株和转化子进行了青霉素发酵及生物活性测定。通过荧光定量PCR(quantitative PCR)对转化子中的Pa-pex11拷贝数进行了相对定量。同时通过透射电镜观察了菌体中微体数量的变化。【结果】实现了灰黄青霉根癌农杆菌介导的遗传转化,获得了在灰黄青霉基因组上插入额外Pa-pex11基因的转化子,在转化子中青霉素产量较野生型菌株提高了1.7倍,电镜照片显示转化子中微体数量明显增加。【结论】通过在基因组中增加Pa-pex11拷贝数能有效地增加灰黄青霉微体的数量,进而提高青霉素的产量。[ Objective] To study the effect of Pa-pex 11 gene on penicillin production in Pencillium aurantiogriseum. [ Method] Based on the conserved domain of pexll from Penicillium chrysogenum, we designed primers pex01 and pex02 to amplify Papexll gene from P. aurantiogriseum. The complete Pa-pexll gene was cloned via thermal asymmetric interlaced PCR （Tail- PCR）. In order to obtain tranformants with additional Pa-pexl 1 gene copy, ATMT （Agrobacterium tumefaciens mediated transformation） was used to transform P. aurantiogriseum. The copy number of Pa-pex 11 was measured by quantitative PCR. Penicillin production of the transformant was analyzed by bioassays and the microbodies were observed by transmission electron microscope （TEM）. [ Results] ATMT was successfully applied in P. aurantiogriseum. Increasing the copy number of Pa-pex 11 gene resulted in a 1.7-fold increase of pencillin production, and the electron microscope graph showed that the number of microbodies increased obviously. [ Conclusion] Our results suggest that increasing the copy number of Pa-pexl 1 can increase the number of micmbodies, and stimulate the penicillin production.

关 键 词：Pa-pex11基因 ATMT 青霉素 微体 

分 类 号：Q933[生物学—微生物学]