基本上不会影响 大家基本上都是这么做的
如果真的要严谨话 可以下列对照实验,这样人家就挑不出你的毛病了
阳性对照:不做任何改动,直接使用带35S的1302.理论上GFP能表达
阴性对照:切除35S启动子,补平缺口,再平端连接.即构建出不带任何启动子驱动下游GFP的阴性对照.观察一下是否能驱动下游GFP表达,理论上GFP不表达.就算能表达,但是是低强度也是可以接受的.
但要注意 pcmbia 系列上的mGFP5 荧光效果不是很好,建议换个增强型的GFP.
"The vectors pCAMBIA1302, pCAMBIA1303, and pCAMBIA1304 are based on pCAMBIA1301 (bacterial kanamycin resistance, plant hygromycin selection, pUC18 polylinker in lacZa) but contain the mgfp5 version of the Aequoria victoria green fluorescent protein (Siemering et al., 1996) either alone (pCAMBIA1302) or in translational fusion with gusA (pCAMBIA1303 has a gusA-mgfp5-His6 fusion, and pCAMBIA1304 has a mgfp5-gusA-His6 fusion).
Unfortunately, Leon Smith reported that analysis of large numbers of transformants of rice and Arabidopsis at CAMBIA showed that the fluorescence produced by the MGFP5 protein was quite faint. CAMBIA staff did then construct similar vectors using the egfp gene available from Clontech, and found results with the EGFP protein were far superior.
However, we are unable to distribute vectors containing this gene. Therefore, we have to recommend that researchers purchase pEGFP from Clontech and use this to construct their own plasmids in any pCAMBIA backbone. "
"considering the poor performance of mgfp5 in pCAMBIA 1302 and others, and since Clonetech did not object on the acquisition of egfp (as per Klaus' messages) from a third party, we can send you Ila the egfp clone that we sent to Klaus and hopefully, it may save you some time, but you may need a mentor with technical skills to help you at work!"
