MicroRNAs (miRNAs) are important for the complex regulation of cell fate decisions and developmental timing.  https://www.selleckchem.com/products/a2ti-2.html  In vivo studies of the contribution of miRNAs during early development are technically challenging due to the limiting cell number. Moreover, many approaches require a miRNA of interest to be defined in assays such as northern blotting, microarray, and qPCR. Therefore, the expression of many miRNAs and their isoforms have not been studied during early development. Here, we demonstrate a protocol for small RNA sequencing of sorted cells from early mouse embryos to enable relatively unbiased profiling of miRNAs in early populations of neural crest cells. We overcome the challenges of low cell input and size selection during library preparation using amplification and gel-based purification. We identify embryonic age as a variable accounting for variation between replicates and stage-matched mouse embryos must be used to accurately profile miRNAs in biological replicates. Our results suggest that this method can be broadly applied to profile the expression of miRNAs from other lineages of cells. In summary, this protocol can be used to study how miRNAs regulate developmental programs in different cell lineages of the early mouse embryo.In this work, we show a detailed engineering route of the first piezoelectric nanostructured epitaxial quartz-based microcantilever. We will explain all the steps in the process starting from the material to the device fabrication. The epitaxial growth of α-quartz film on SOI (100) substrate starts with the preparation of a strontium doped silica sol-gel and continues with the deposition of this gel into the SOI substrate in a thin film form using the dip-coating technique under atmospheric conditions at room temperature. Before crystallization of the gel film, nanostructuration is performed onto the film surface by nanoimprint lithography (NIL). Epitaxial film growth is reached at 1000 °C, inducing a perfect crystallization of the patterned gel film. Fabrication of quartz crystal cantilever devices is a four-step process based on microfabrication techniques. The process starts with shaping the quartz surface, and then metal deposition for electrodes follows it. After removing the silicone, the cantilever is released from SOI substrate eliminating SiO2 between silicon and quartz. The device performance is analyzed by non-contact laser vibrometer (LDV) and atomic force microscopy (AFM). Among the different cantilever's dimensions included in the fabricated chip, the nanostructured cantilever analyzed in this work exhibited a dimension of 40 µm large and 100 µm long and was fabricated with a 600 nm thick patterned quartz layer (nanopillar diameter and separation distance of 400 nm and 1 µm, respectively) epitaxially grown on a 2 µm thick Si device layer. The measured resonance frequency was 267 kHz and the estimated quality factor, Q, of the whole mechanical structure was Q ~ 398 under low vacuum conditions. We observed the voltage-dependent linear displacement of cantilever with both techniques (i.e., AFM contact measurement and LDV). Therefore, proving that these devices can be activated through the indirect piezoelectric effect.Nuclear magnetic resonance (NMR) spectroscopy represents an important technique to understand the structure and bonding environments of molecules. There exists a drive to characterize materials under conditions relevant to the chemical process of interest. To address this, in situ high-temperature, high-pressure MAS NMR methods have been developed to enable the observation of chemical interactions over a range of pressures (vacuum to several hundred bar) and temperatures (well below 0 °C to 250 °C). Further, the chemical identity of the samples can be comprised of solids, liquids, and gases or mixtures of the three. The method incorporates all-zirconia NMR rotors (sample holder for MAS NMR) which can be sealed using a threaded cap to compress an O-ring. This rotor exhibits great chemical resistance, temperature compatibility, low NMR background, and can withstand high pressures. These combined factors enable it to be utilized in a wide range of system combinations, which in turn permit its use in diverse fields as carbon sequestration, catalysis, material science, geochemistry, and biology. The flexibility of this technique makes it an attractive option for scientists from numerous disciplines.Quality control in botanical products begins with the raw material supply. Traditionally, botanical identification is performed through morphological assessment and chemical analytical methods. However, the lack of availability of botanists, especially in recent years, coupled with the need to enhance quality control to combat the stresses on the supply chain brought by increasing consumer demand and climate change, necessitates alternative approaches. The goal of this protocol is to facilitate botanical species identification using a portable qPCR system on the field or in any setting, where access to laboratory equipment and expertise is limited. Target DNA is amplified using dye-based qPCR, with DNA extracted from botanical reference materials serving as a positive control. The target DNA is identified by its specific amplification and matching its melting peak against the positive control. A detailed description of the steps and parameters, from hands-on field sample collection, to DNA extraction, PCR amplification, followed by data interpretation, has been included to ensure that readers can replicate this protocol. The results produced align with traditional laboratory botanical identification methods. The protocol is easy to perform and cost-effective, enabling quality testing on raw materials as close to the point of origin of the supply chain as possible.Testicular organoids provide a tool for studying testicular development, spermatogenesis, and endocrinology in vitro. Several methods have been developed in order to create testicular organoids. Many of these methods rely upon extracellular matrix (ECM) to promote de novo tissue assembly, however, there are differences between methods in terms of biomimetic morphology and function of tissues. Moreover, there are few direct comparisons of published methods. Here, a direct comparison is made by studying differences in organoid generation protocols, with provided outcomes. Four archetypal generation methods (1) 2D ECM-free, (2) 2D ECM, (3) 3D ECM-free, and (4) 3D ECM culture are described. Three primary benchmarks were used to assess the testicular organoid generation. These are cellular self-assembly, inclusion of major cell types (Sertoli, Leydig, germ, and peritubular cells), and appropriately compartmentalized tissue architecture. Of the four environments tested, 2D ECM and 3D ECM-free cultures generated organoids with internal morphologies most similar to native testes, including the de novo compartmentalization of tubular versus interstitial cell types, the development of tubule-like-structures, and an established long-term endocrine function.