Isolation of two highly active soybean (Glycine max (L.) Merr.) promoters and their characterization using a new automated image collection and analysis system
Isolation of two highly active soybean (Glycine max (L.) Merr.) promoters and their characterization using a new automated image collection and analysis system
复制标题
DOI：
10.1007/s00299-007-0359-y
复制
发表时间：
2007-09-01
影响因子：
6.2
通讯作者：
Finer, John J.
中科院分区：
文献类型：
--
作者：
Chiera, Joseph M.;Bouchard, Robert A.;Finer, John J.
利用一种新型的自动图像采集和分析系统，对两个新的大豆品种(Glyine max(L.)(Merr.)以花椰菜花叶病毒35S(CaMV35S)启动子为表达标准。为了进行表达比较，将大豆多聚泛素(Gmubi)启动子、类似大豆热休克蛋白90(GmHSP90L)启动子和CaMV35S启动子的不同排列置于绿色荧光蛋白(GFP)基因的上游。用基因枪法将DNA载体导入萌发的菜豆(Phaseolus Lunatus L.)离体子叶。种子，放在培养皿中，用于自动图像捕获和图像分析。这个自动化系统可以随着时间的推移监测和量化同一组织中GFP基因的表达。内含子完整的Gmubi启动子的表达强度最高，是CaMV35S启动子的5倍以上。当从Gmubi启动子中去除内含子区域时，GFP的表达降低，但仍比CaMV35S启动子的表达高两倍以上。大豆GmHSP90L全长启动子比CaMV35S启动子强4倍。GmHSP90L启动子的截短导致启动子强度的逐步降低，这似乎与调控元件的移除相对应。自动图像捕获和分析使我们能够快速高效地评估这些新的推广者。
A novel automated image collection and analysis system was used to compare two new soybean (Glycine max (L.) Merr.) promoters with the cauliflower mosaic virus 35S (CaMV35S) promoter, which was used as an expression standard. For expression comparisons, various permutations of a soybean polyubiquitin (Gmubi) promoter, a soybean heat shock protein 90-like (GmHSP90L) promoter and the CaMV35S promoter were placed upstream of a green fluorescent protein (gfp) gene. DNA constructs were introduced via particle bombardment into excised cotyledons of germinating lima bean (Phaseolus lunatus L.) seeds, which were arranged in Petri dishes for automated image capture and image analysis. The automated system allowed monitoring and quantification of gfp gene expression in the same piece of tissue over time. The Gmubi promoter, with its intronic region intact, showed the highest expression that was over five times stronger than the CaMV35S promoter. When an intronic region was removed from the Gmubi promoter, GFP expression was reduced, but was still over two times greater than with the CaMV35S promoter. The full-length soybean GmHSP90L promoter was four times stronger than the CaMV35S promoter. Truncation of the GmHSP90L promoter resulted in stepwise decreases in promoter strength, which appear to correspond to removal of regulatory elements. Automated image capture and analysis allowed the rapid and efficient evaluation of these new promoters.