Milner Hayley, Nowak Scott J
Department of Molecular and Cellular Biology, Kennesaw State University, United States.
MethodsX. 2020 Nov 5;7:101130. doi: 10.1016/j.mex.2020.101130. eCollection 2020.
is a powerful model organism in which to address the genetics of cardiac patterning and heart development. This system allows the pairing of live imaging with the myriad available genetic and transgenic techniques to not only identify the genes that are critical for heart development, but to assess their impact on heart function in living organisms. There are several described methods to assess cardiac function in However, these approaches are restricted to imaging of mid- to late-instar larval and adult hearts. This technical hurdle therefore does not allow for the recording and analysis of cardiac function in embryos bearing strong mutations that do not hatch into larvae. Our technical innovation lies in transgenically labeling the cells of the heart and using line scan-based confocal imaging to repeatedly image the walls of the heart. By plotting this line scan as a kymograph, heart contractions can be visualized and assayed, thereby allowing for quantification of physiological defects. This method can be used to obtain physiological data from known mutations that affect cardiac development yet are incapable of hatching into larvae for conventional analysis.•Use transgenic methods to label heart proper walls•Use high-speed line scanning to capture position of heart proper walls•Create X vs. time plot to visualize and quantify contractions over imaging period.
是一种强大的模式生物，可用于研究心脏模式形成和心脏发育的遗传学。该系统允许将活体成像与众多可用的遗传和转基因技术相结合，不仅可以识别对心脏发育至关重要的基因，还可以评估它们对活生物体心脏功能的影响。有几种已描述的方法可用于评估心脏功能。然而，这些方法仅限于对中晚期幼虫和成虫心脏进行成像。因此，这一技术障碍不允许对携带不孵化成幼虫的强突变胚胎的心脏功能进行记录和分析。我们的技术创新在于通过转基因标记心脏细胞，并使用基于线扫描的共聚焦成像反复对心脏壁进行成像。通过将此线扫描绘制为波形图，可以可视化和分析心脏收缩，从而实现生理缺陷的量化。这种方法可用于从已知的影响心脏发育但无法孵化成幼虫进行传统分析的突变中获取生理数据。•使用转基因方法标记心脏固有壁•使用高速线扫描捕获心脏固有壁的位置•创建X与时间图以在成像期间可视化和量化收缩。
