Beanirwin9409

Single-molecule RNA fluorescence in situ hybridization (smFISH) allows subcellular visualization, localization, and quantification of endogenous RNA molecules in fixed cells. The spatial and intensity information of each RNA can be used to distinguish mature from nascent transcripts inside each cell, revealing both past and instantaneous transcriptional activity. Here, we describe an optimized protocol for smFISH in Saccharomyces cerevisiae with optimized lyticase digestion time and hybrization steps for more homogenous results. For complete details on the use and execution of this protocol, please refer to Donovan et al. (2019).Neonatal mouse cochlear duct cells can proliferate and grow in vitro into inner ear organoids. Distinctive cochlear duct cell types have different organoid formation capacities. Here, we provide a flow cytometric cell-sorting method that allows the subsequent culture of individual cochlear cell populations. For the efficient culture of the sorted cells, we provide protocols for growing free-floating inner ear organoids, the adherence of organoids to a substrate, and the expansion of organoid-derived inner ear colonies. For complete details on the use and execution of this protocol, please refer to Kubota et al. (2021).Co-immunoprecipitation (co-IP) of protein complexes from cell lysates is widely used to study protein-protein interactions. However, establishing robust co-IP assays often involves considerable optimization. Moreover, co-IP results are frequently presented in non-quantitative ways. This protocol presents an optimized co-IP workflow with an analysis based on semi-quantitative immunoblot densitometry to increase reliability and reproducibility. For complete details on the use and execution of this protocol, please refer to Burckhardt et al. Deferoxamine (2021).In S. cerevisiae, we identified rhomboid pseudoprotease Dfm1 as the major mediator for removing or retrotranslocating misfolded membrane substrates from the ER (endoplasmic reticulum). Long-standing challenges with rapid suppression of dfm1-null cells have limited the biochemical study of Dfm1's role in ER protein quality control. Here, we provide a protocol for the generation and handling of dfm1-null cells and procedures for studying normal vs. suppressive alternative retrotranslocation pathways. Our methods can be utilized to study other components involved in retrotranslocation. For complete information on the generation and use of this protocol, please refer to Neal et al. (2017, 2018); Neal et al. (2019); Neal et al. (2020).In the first 6 months of 2020, the COVID-19 pandemic forced numerous universities across the globe to quickly transfer all their courses online, a response known as Emergency Remote Teaching. Courses initially designed for face to face delivery had to be quickly adapted to a new online format. In this paper, we study the perception of the transition to remote teaching in a group of computer science students. Despite the advantage given by an average higher computer literacy, the results of this study suggest that students found the transition challenging, especially regarding tasks such as asking questions during video lectures and interacting with instructors. The transition seems to have had a greater impact on students of lower level courses. Differences were found also across race and residence status (but not gender). The initial format of the course was also relevant students fared better if their course relied on online tools before the transition.The digital world has a wealth of data, such as internet of things (IoT) data, business data, health data, mobile data, urban data, security data, and many more, in the current age of the Fourth Industrial Revolution (Industry 4.0 or 4IR). Extracting knowledge or useful insights from these data can be used for smart decision-making in various applications domains. In the area of data science, advanced analytics methods including machine learning modeling can provide actionable insights or deeper knowledge about data, which makes the computing process automatic and smart. In this paper, we present a comprehensive view on "Data Science" including various types of advanced analytics methods that can be applied to enhance the intelligence and capabilities of an application through smart decision-making in different scenarios. We also discuss and summarize ten potential real-world application domains including business, healthcare, cybersecurity, urban and rural data science, and so on by taking into account data-driven smart computing and decision making. Based on this, we finally highlight the challenges and potential research directions within the scope of our study. Overall, this paper aims to serve as a reference point on data science and advanced analytics to the researchers and decision-makers as well as application developers, particularly from the data-driven solution point of view for real-world problems.

Well-defined germ-line mutations in the

gene are associated with syndromic multiple basal cell carcinomas (BCCs). Here, we used whole exome sequencing (WES) to identify the role of patched-1 in patients with multiple, unusually large BCCs.

A 72-year old patient presenting with numerous BCCs progressing to large ulcerating lesions was enrolled. WES was used to identify the pathogenic gene locus.

Genetic work-up by WES identified a homozygous

nonsense mutation in the tumor tissue but not present in her blood cells or in non-lesional skin. In addition, heterozygous missense mutations were identified in three cancer-associated genes (

, and

) in blood cells as well as in lesional and non-lesional skin. We also tested systemic immune therapy as a potentially beneficial approach to treat patients with numerous large BCCs on scatted areas of involvement. A rapid and sustained response to nivolumab was noted, suggesting that it is an efficacious drug for long-term therapeutic outcome.

,

, and

may potentially contribute to the synergistic oncogene driven malignant transformation manifesting as multiple, unusually large BCCs.

PTCH1, EPHB2, RET, and GALNT12 may potentially contribute to the synergistic oncogene driven malignant transformation manifesting as multiple, unusually large BCCs.