单子叶植物不同Ppc基因构建对烟草转化幼苗生长的影响

Different Monocotyledonous of Ppc Genes on the Growth of Transformed Tobacco Seedlings

为比较单子叶植物2种不同形式(c DNA和DNA)的磷酸烯醇式丙酮酸羧化酶(phosphoenolpyruvate carboxylase,PEPC)基因(Ppc)在双子叶植物中的表达效果,本研究将单子叶植物稗草(Echinochloa crusgalli)Ppc基因的c DNA和玉米(Zea mays)Ppc基因的DNA全长通过农杆菌介导对双子叶植物烟草(Nicotiana tabacum)进行了遗传转化。GUS组织化学染色、PCR、RT-PCR检测结果表明,2种形式的Ppc基因均转入了烟草中;观察分化幼苗生长发育过程发现:转Ppc基因c DNA的烟草分化苗生长发育正常,而转Ppc基因DNA的分化幼苗在培养过程中叶片白化现象严重,未完成正常生长发育过程,可能是由于完整基因DNA在异源细胞中转录起始位点不正确或拼接错误而导致基因表达异常。由此推测在遗传距离较远的物种间转移DNA基因序列全长可能会降低正常表达率。净光合速率(Pn)的测定结果表明,大部分转稗草Ppc基因烟草的Pn高于对照,结果初步证明单子叶植物稗草的根型PEPC酶对烟草的光合作用具有一定的调节作用。

To compare two different forms （cDNA in monocotyledonous plants Echinochloa, DNA in monocotyledon Zea mays） ofphosphoenolpyruvate carboxylase （PEPC） gene （Ppc） on transformed dicotyledonous plant growth, the Ppc genes wereAgrobactirium-transfected into dicotyledon tobacco. Transformed leaf discs and differentiated seedling leaves were verified withGUS histochemistry, PCR, and RT-PCR. It was found that tobaccos transformed with barnyardgrass cDNA grew better than thecomplete maize DNA Ppc gene. The latter tobacco plants showed lower regeneration efficiency, leaves turned yellow and appearedwilted, possibly due to abnormal chloroplasts, or the complete maize Ppc gene was not expressed correctly in tobacco due toincorrect transcription initiation or incorrect splicing. Therefore to transfer DNA gene between genetically-distant species lowernormal expression rate. Net photosynthetic rate （Pn） of transgenic tobacco plants transformed with barnyardgrass cDNA was higherthan in untransformed tobacco, indicating that over-expressing Echinochloa root-Ppc gene improved tobacco photosynthesis.