Light Sheet Tomography (LST) for in situ imaging of plant roots

Abstract

The production of crops capable of efficient nutrient use is essential for addressing the problem of global food security. The ability of a plant's root system to interact with the soil micro-environment determines how effectively it can extract water and nutrients. In order to assess this ability and develop the fast and cost effective phenotyping techniques which are needed to establish efficient root systems, in situ imaging in soil is required. To date this has not been possible due to the high density of scatterers and absorbers in soil or because other growth substrates do not sufficiently model the heterogeneity of a soil's microenvironment. We present here a new form of light sheet imaging with novel transparent soil containing refractive index matched particles. This imaging method does not rely on fluorescence, but relies solely on scattering from root material. We term this form of imaging Light Sheet Tomography (LST). We have tested LST on a range of materials and plant roots in transparent soil and gel. Due to the low density of root structures, i.e. relatively large spaces between adjacent roots, long-term monitoring of lettuce root development in situ with subsequent quantitative analysis was achieved.

A scanner system for high-resolution quantification of variation in root growth dynamics of Brassica rapa genotypes

“…Biospeckle laser imaging, a more recent technique, provides new opportunities to screen for functional traits without the use of a dye (Ribeiro et al , 2014). Finally, transparent substrates allow 3D descriptions of RSAs using a range of techniques such as laser scanning (Fang et al , 2009), optical (Iyer-Pascuzzi et al , 2010; Clark et al , 2011), or light sheet tomography (Yang et al , 2013). …”

Light-sheet microscopy: a tutorial

“…Optical scattering collected from unlabeled biological tissue would provide valuable structural information that can Tutorial originate from different biological compartments and interfaces, including tissue boundaries, cells, organelles, or macromolecular complexes. In spite of this, elastic scattering has barely been exploited as a contrast mechanism to generate LSFM images [114]; please see Section 6.1d for specific details on elastic scattering LSFM implementations.…”

“…On the other hand, when the scattering is elastic, it would give information about the structure of the sample at different spatial scale levels, as discussed in Section 2.3, and with minimum light dosage deposited onto the sample. LS elastic scattering microscopy has been implemented and demonstrated for plant root phenotyping embedded in transparent soils and quantitative long-term monitoring of lettuce roots [114]. Nevertheless, the contrast and the resolution of the obtained images were seen to be highly dependent on the turbidity and refractive index heterogeneity of both the sample and the embedding substrate.…”

Gaussian vs. Bessel light-sheets: performance analysis in live large sample imaging

“…Lightsheet imaging is no exception to this [12]. In fact, it is possible to combine scattering imaging along with fluorescence imaging to get dual contrast images of chick embryos.…”