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1. Single-cell Iso-Sequencing enables rapid genome annotation for scRNAseq analysis

   https://doi.org/10.1093/genetics/iyac017  

   Healey, Hope_M; Bassham, Susan; Cresko, William_A; Sanchez_Alvarado, ed., A. (February 2022, Genetics)

   Abstract Single-cell RNA sequencing is a powerful technique that continues to expand across various biological applications. However, incomplete 3′-UTR annotations can impede single-cell analysis resulting in genes that are partially or completely uncounted. Performing single-cell RNA sequencing with incomplete 3′-UTR annotations can hinder the identification of cell identities and gene expression patterns and lead to erroneous biological inferences. We demonstrate that performing single-cell isoform sequencing in tandem with single-cell RNA sequencing can rapidly improve 3′-UTR annotations. Using threespine stickleback fish (Gasterosteus aculeatus), we show that gene models resulting from a minimal embryonic single-cell isoform sequencing dataset retained 26.1% greater single-cell RNA sequencing reads than gene models from Ensembl alone. Furthermore, pooling our single-cell sequencing isoforms with a previously published adult bulk Iso-Seq dataset from stickleback, and merging the annotation with the Ensembl gene models, resulted in a marginal improvement (+0.8%) over the single-cell isoform sequencing only dataset. In addition, isoforms identified by single-cell isoform sequencing included thousands of new splicing variants. The improved gene models obtained using single-cell isoform sequencing led to successful identification of cell types and increased the reads identified of many genes in our single-cell RNA sequencing stickleback dataset. Our work illuminates single-cell isoform sequencing as a cost-effective and efficient mechanism to rapidly annotate genomes for single-cell RNA sequencing. 

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2. Thermo-mechanically responsive crystalline organic cantilever

   https://doi.org/10.1039/C7CC04346E  

   Dharmarwardana, Madushani; Welch, Raymond P.; Kwon, Sunah; Nguyen, Victoria K.; McCandless, Gregory T.; Omary, Mohammad A.; Gassensmith, Jeremiah J. (January 2017, Chem. Commun.)

   We report thermo-mechanically responsive and thermochromic behavior in the single crystalline organic semiconductor butoxyphenyl N -substituted naphthalene diimide (BNDI). We show that initially monoclinic single crystals of BNDI undergo a single-crystal to single-crystal transition to a triclinic phase. This transition accompanies large changes in the crystal packing, a visible decrease in crystal size, reversible thermochromic behavior, and motion including bending, jumping, and splitting. We have shown that by fixing single crystals to a surface, we can harness the energy of the phase transition to create a single crystal cantilever capable of lifting weights with masses nigh two orders of magnitude heavier than the single crystal itself. 

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3. Real-time 3D single molecule tracking

   https://doi.org/10.1038/s41467-020-17444-6  

   Hou, Shangguo; Exell, Jack; Welsher, Kevin (July 2020, Nature Communications)

   Abstract To date, single molecule studies have been reliant on tethering or confinement to achieve long duration and high temporal resolution measurements. Here, we present a 3D single-molecule active real-time tracking method (3D-SMART) which is capable of locking on to single fluorophores in solution for minutes at a time with photon limited temporal resolution. As a demonstration, 3D-SMART is applied to actively track single Atto 647 N fluorophores in 90% glycerol solution with an average duration of ~16 s at count rates of ~10 kHz. Active feedback tracking is further applied to single proteins and nucleic acids, directly measuring the diffusion of various lengths (99 to 1385 bp) of single DNA molecules at rates up to 10 µm²/s. In addition, 3D-SMART is able to quantify the occupancy of single Spinach2 RNA aptamers and capture active transcription on single freely diffusing DNA. 3D-SMART represents a critical step towards the untethering of single molecule spectroscopy. 

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4. Electrospray deposition for single nanoparticle studies

   https://doi.org/10.1039/d1ay01295a  

   Jagdale, Gargi S.; Choi, Myung-Hoon; Siepser, Natasha P.; Jeong, Soojin; Wang, Yi; Skalla, Rebecca X.; Huang, Kaixiang; Ye, Xingchen; Baker, Lane A. (August 2021, Analytical Methods)

   null (Ed.)

   Single entity electrochemical (SEE) studies that can probe activities and heterogeneity in activities at nanoscale require samples that contain single and isolated particles. Single, isolated nanoparticles are achieved here with electrospray deposition of colloidal nanoparticle solutions, with simple instrumentation. Role of three electrospray (ES) parameters, viz. spray distance (emitter tip-to-substrate distance), ES current and emitter tip diameter, in the ES deposition of single Au nano-octahedra (Au ODs) is examined. The ES deposition of single, isolated Au ODs are analyzed in terms of percentage of single NPs and local surface density of deposition. The local surface density of ES deposition of single Au ODs was found to increase with decrease in spray distance and emitter tip diameter, and increase in ES current. While the percentage of single particle ES deposition increased with increase in spray distance and decrease in emitter tip size. No significant change in the single Au ODs ES deposition percentage was observed with change in ES current values included in this study. The most favourable conditions in the ES deposition of Au ODs in this study resulted in the local surface density of 0.26 ± 0.05 single particles per μm 2 and observation of 96.3% single Au OD deposition. 

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5. Leveraging Single-Cell Populations to Uncover the Genetic Basis of Complex Traits

   https://doi.org/10.1146/annurev-genet-022123-110824  

   Minow, Mark A.A.; Marand, Alexandre P.; Schmitz, Robert J. (November 2023, Annual Review of Genetics)

   The ease and throughput of single-cell genomics have steadily improved, and its current trajectory suggests that surveying single-cell populations will become routine. We discuss the merger of quantitative genetics with single-cell genomics and emphasize how this synergizes with advantages intrinsic to plants. Single-cell population genomics provides increased detection resolution when mapping variants that control molecular traits, including gene expression or chromatin accessibility. Additionally, single-cell population genomics reveals the cell types in which variants act and, when combined with organism-level phenotype measurements, unveils which cellular contexts impact higher-order traits. Emerging technologies, notably multiomics, can facilitate the measurement of both genetic changes and genomic traits in single cells, enabling single-cell genetic experiments. The implementation of single-cell genetics will advance the investigation of the genetic architecture of complex molecular traits and provide new experimental paradigms to study eukaryotic genetics. 

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