Spatiotemporal mapping of gene expression to reveal mechanisms of cellular differentiation in E. coli biofilms

在第二个财政年度，我们在多个方面取得了进展。1）我们开始并取得了进展，通过在不同径向位置对生物膜进行采样，然后进行蛋白质组分析，对蛋白质表达进行空间分析。2）我们进一步表征了不同条件下选定启动子活性的表达。为了使荧光结果标准化，我们一直在测试一种二级荧光蛋白，其表达是组成型的。3）我们一直在表征葡萄糖添加对生物膜发育的影响，我们的结果表明，意料之外的代谢失衡可能在该过程中很重要。4）我们开发了基于Raspberry Pi计算机和相机的成像系统，以收集时间-除了亮场之外，还可以在双荧光模式下拍摄延时图像。

During the second fiscal year we have progressed on multiple fronts.1) We started and progressed with spatial analysis of protein expression by sampling biofilms at different radial locations followed by ongoing proteome analysis.2) We characterized further expression of selected promoters activity under different conditions. To normalize the fluorescence results we have been testing a secondary fluorescent protein whose expression is constitutive.3) We have been characterizing the effect of glucose addition on biofilm development and our results suggest that an unsuspected metabolic imbalance may be important in the process.4) We have developed imaging systems based on Raspberry Pi computers and cameras to collect time-lapse images in dual fluorescence mode in addition to bright field.