Synchrotron Based X-ray Microtomography Reveals Cellular Morphological Features of Developing Wheat Grain  
[PERSON]  
1 INRAE, UR BIA, 44316 Nantes, France; [EMAIL_ADDRESS] (C.A. ); [EMAIL_ADDRESS] (F.G.); [EMAIL_ADDRESS] (T.D.Q.L. ); [EMAIL_ADDRESS] (L.P.); [PERSON]
2 INRAE, PROBE Research Infrastructure, BIIBS Facility, 44316 Nantes, France 2  
[PERSON]  
45  
[PERSON]  
1 INRAE, UR BIA, 44316 Nantes, France; [EMAIL_ADDRESS] (C.A. ); [EMAIL_ADDRESS] (F.G.); [EMAIL_ADDRESS] (T.D.Q.L. ); [EMAIL_ADDRESS] (L.P.); [PERSON]
6  
Amber King  
45  
Thang Duong Quoc Le  
45  
[PERSON]  
45  
[PERSON]  
12 INRAE, UR BIA, 44316 Nantes, France; [EMAIL_ADDRESS] (C.A. ); [EMAIL_ADDRESS] (F.G.); [EMAIL_ADDRESS] (T.D.Q.L. ); [EMAIL_ADDRESS] (L.P.); [EMAIL_ADDRESS] (A.-L.C.-B.) 6  
[PERSON]  
6,7 pt  
main parts of the grain. The grain development is initiated by ovule fertilization. However, it is highly dependent on the prior ovary development [4]. Grain development is classically divided into three phases: early-(lag-) phase (from 10 to 15 days after anthesis), filling phase, and maturation phase [5]. At anthesis, the wheat ovary consists of ovary tissues surrounding an ovule. Following fertilization, the ovary epidermis and parenchyma give the grain pericarp tissues (epicarp or pericarp epidermis and mesocarp, or pericarp parenchyma), while the endosperm and embryo develop. In the very early stages of development, the cereal grain is composed mostly of outer tissues: pericarp and integuments. The endosperm forms from free nuclei, then cellularization takes place followed by cell division, and cell differentiation into starch endosperm, aleurone, and transfer cells. At the end of the early phase of development, the wheat grain reaches its final length. Between 15 and 35-40 days after anthesis, the endosperm fills with storage compounds (mainly starch and proteins). During this phase, the outer tissues undergo drastic changes. In particular, the mesocarp parenchyma cells undergo programmed cell death [6]. At the end of the filling phase, the grain enters the maturation phase and desiccates because of water loss. During this early development, the shape of the wheat grain changes from a triangular shape to an ellipsoid [7], and a crease and two lobes appear. The study of plant tissues anatomy traditionally involves optical microscopy of stained cross-sections and this is the case for the wheat grain [8; 9; 10]. Early observations of wheat grain tissues were conducted using transmission or scanning electron microscopy on sections or dissected tissues [11; 12; 13]. However, as grain development is not spatially uniform [14], understanding the development or the physiology of the grain needs to investigate its anatomy in three dimensions. Several possibilities are available to study entirely the 3D anatomy of a plant organ. The acquisition by optical or electron microscopy of a large number of serial sections, followed by a registration process, can lead to the reconstruction of the 3D anatomy [15; 16; 17; 18]. Magnetic resonance imaging is a non-destructive imaging technique that can be used to analyze both the anatomy and the functions of plant tissues at the organ scale [19; 20; 21]. The resolution is however limited to a few dozens of microns, making it difficult to investigate thin tissues or cell organization. X-ray computed tomography is a popular method for the non-destructive investigation of the 3D architecture of biological specimens, without requiring staining, sectioning, or inclusion. The high resolution reached by computed tomography makes it a method of choice for the investigation of plant organs [22; 23; 24; 25; 26]. X-ray computed tomography makes use of many X-ray measurements taken from different angles to produce projection images. The set of projections can be processed to generate a 3D image of the whole specimen, allowing observation through virtual slices along arbitrary angles. Dry specimens present high contrast to X-rays, and many impressive 3D reconstructions have been performed on seeds [27; 28].