Deciphering single-cell 3D chromatin structure using scCTG.

Sequencing-based Hi-C technology has been widely used to study the three-dimensional structure of chromatin. More recently, the development of single-cell Hi-C technology has enabled the study of chromatin structural variations between individual cells. However, single-cell Hi-C data are often highly sparse, necessitating the use of imputation algorithms to address insufficient sampling.

BC-Assisted Paper Spray Ionization Mass Spectrometry.

A novel inorganic nonmetallic material boron carbide (BC) was applied in paper spray ionization to analyze organic molecules in both negative and positive modes. The utilization of BC has led to a notable enhancement in signal responses for various molecules, including bisphenols and drugs, by approximately two to four times. The limit of detection (LOD) of bisphenol AF and nilotinib standard solutions can reach 1 and 0.

Complex structure and activation mechanism of arginine kinase McsB by McsA.

Protein phosphorylation is a pivotal post-translational modification modulating various cellular processes. In Gram-positive bacteria, the protein arginine kinase McsB, along with its activator McsA, has a key role in labeling misfolded and damaged proteins during stress. However, the activation mechanism of McsB by McsA remains elusive.

Modulation of the microhomology-mediated end joining pathway suppresses large deletions and enhances homology-directed repair following CRISPR-Cas9-induced DNA breaks.

Background: CRISPR-Cas9 genome editing often induces unintended, large genomic rearrangements, posing potential safety risks. However, there are no methods for mitigating these risks.

Results: Using long-read individual-molecule sequencing (IDMseq), we found the microhomology-mediated end joining (MMEJ) DNA repair pathway plays a predominant role in Cas9-induced large deletions (LDs).

Fully synthetic phosphorylated Tau181, Tau217, and Tau231 calibrators for Alzheimer's disease diagnosis.

Background: The calibrator in immunoassay plays an essential role in diagnosing Alzheimer's disease (AD). Presently, the most well-studied biomarkers for AD diagnosis are three phosphorylated Tau (p-Tau): p-Tau231, p-Tau217, and p-Tau181. Glycogen synthase-3beta (GSK3β)-phosphorated Tau-441 is the most commonly used calibrator for p-Tau immunoassays.

Rapid and signal crowdedness-robust in situ sequencing through hybrid block coding.

Spatial transcriptomics technology has revolutionized our understanding of cell types and tissue organization, opening possibilities for researchers to explore transcript distributions at subcellular levels. However, existing methods have limitations in resolution, sensitivity, or speed. To overcome these challenges, we introduce SPRINTseq (Spatially Resolved and signal-diluted Next-generation Targeted sequencing), an innovative in situ sequencing strategy that combines hybrid block coding and molecular dilution strategies.

Multi-omics for COVID-19: driving development of therapeutics and vaccines.

The ongoing COVID-19 pandemic caused by SARS-CoV-2 has raised global concern for public health and economy. The development of therapeutics and vaccines to combat this virus is continuously progressing. Multi-omics approaches, including genomics, transcriptomics, proteomics, metabolomics, epigenomics and metallomics, have helped understand the structural and molecular features of the virus, thereby assisting in the design of potential therapeutics and accelerating vaccine development for COVID-19.

Ancient DNA reveals genetic admixture in China during tiger evolution.

The tiger (Panthera tigris) is a charismatic megafauna species that originated and diversified in Asia and probably experienced population contraction and expansion during the Pleistocene, resulting in low genetic diversity of modern tigers. However, little is known about patterns of genomic diversity in ancient populations. Here we generated whole-genome sequences from ancient or historical (100-10,000 yr old) specimens collected across mainland Asia, including a 10,600-yr-old Russian Far East specimen (RUSA21, 8× coverage) plus six ancient mitogenomes, 14 South China tigers (0.

Hydrogel-Based Multi-enzymatic System for Biosynthesis.

Biosynthesis involving multi-enzymatic reactions is usually an efficient and economic method to produce plentiful important molecules. To increase the product yield in biosynthesis, the involved enzymes can be immobilized to carriers for enhancing enzyme stability, increasing synthesis efficiency and improving enzyme recyclability. Hydrogels with three-dimensional porous structures and versatile functional groups are promising carriers for enzyme immobilization.

Tracking the disulfide rearrangement of heated lactoglobulin by matrix-assisted laser desorption/ionization-in-source decay top-down analysis.

Disulfide bond rearrangement is a common occurrence during protein analysis or treatment. A convenient and rapid method has been developed to investigate heat-induced disulfide rearrangement of lactoglobulin using matrix-assisted laser desorption/ionization-in-source decay (MALDI-ISD) technology. By analyzing heated lactoglobulin in reflectron and linear mode, we demonstrated that cysteines C66 and C160 exist as free residues other than linked ones in some protein isomers.

Stochastic Monte Carlo Model for Simulating the Dynamic Liquid-Liquid Phase Separation in Bacterial Cells.

There is growing evidence showing that many critical biological processes are driven by biomolecule condensates through liquid-liquid phase separation (LLPS). Although the qualitative observation and description of LLPS have been well documented, quantitative simulations of the time-dependent progression of LLPS in live cells are generally lacking. In this work, we build a stochastic Monte Carlo model to simulate the dynamic LLPS process during the formation of bacterial aggresomes.

Large-scale high-throughput 3D culture, imaging, and analysis of cell spheroids using microchip-enhanced light-sheet microscopy.

Light sheet microscopy combined with a microchip is an emerging tool in biomedical research that notably improves efficiency. However, microchip-enhanced light-sheet microscopy is limited by noticeable aberrations induced by the complex refractive indices in the chip. Herein, we report a droplet microchip that is specifically engineered to be capable of large-scale culture of 3D spheroids (over 600 samples per chip) and has a polymer index matched to water (difference <1%).

Quantitative haplotype-resolved analysis of mitochondrial DNA heteroplasmy in Human single oocytes, blastoids, and pluripotent stem cells.

Maternal mitochondria are the sole source of mtDNA for every cell of the offspring. Heteroplasmic mtDNA mutations inherited from the oocyte are a common cause of metabolic diseases and associated with late-onset diseases. However, the origin and dynamics of mtDNA heteroplasmy remain unclear.

Single-cell individual full-length mtDNA sequencing by iMiGseq uncovers unexpected heteroplasmy shifts in mtDNA editing.

The ontogeny and dynamics of mtDNA heteroplasmy remain unclear due to limitations of current mtDNA sequencing methods. We developed individual Mitochondrial Genome sequencing (iMiGseq) of full-length mtDNA for ultra-sensitive variant detection, complete haplotyping, and unbiased evaluation of heteroplasmy levels, all at the individual mtDNA molecule level. iMiGseq uncovered unappreciated levels of heteroplasmic variants in single cells well below the conventional NGS detection limit and provided accurate quantitation of heteroplasmy level.

Assessment of microbiota in the gut and upper respiratory tract associated with SARS-CoV-2 infection.

Background: The human microbiome plays an important role in modulating the host metabolism and immune system. Connections and interactions have been found between the microbiome of the gut and oral pharynx in the context of SARS-CoV-2 and other viral infections; hence, to broaden our understanding of host-viral responses in general and to deepen our knowledge of COVID-19, we performed a large-scale, systematic evaluation of the effect of SARS-CoV-2 infection on human microbiota in patients with varying disease severity.

Results: We processed 521 samples from 203 COVID-19 patients with varying disease severity and 94 samples from 31 healthy donors, consisting of 213 pharyngeal swabs, 250 sputa, and 152 fecal samples, and obtained meta-transcriptomes as well as SARS-CoV-2 sequences from each sample.

Surface modification for improving immunoassay sensitivity.

Immunoassays are widely performed in many fields such as biomarker discovery, proteomics, drug development, and clinical diagnosis. There is a growing need for high sensitivity of immunoassays to detect low abundance analytes. As a result, great effort has been made to improve the quality of surfaces, on which the immunoassay is performed.

Single-cell RNA-seq reveals early heterogeneity during aging in yeast.

The budding yeast Saccharomyces cerevisiae (S. cerevisiae) has relatively short lifespan and is genetically tractable, making it a widely used model organism in aging research. Here, we carried out a systematic and quantitative investigation of yeast aging with single-cell resolution through transcriptomic sequencing.

Rapid and sensitive single-cell RNA sequencing with SHERRY2.

Background: Prevalent single-cell transcriptomic profiling (scRNA-seq) methods are mainly based on the synthesis and enrichment of full-length double-stranded complementary DNA. These approaches are challenging to generate accurate quantification of transcripts when their abundance is low or their full-length amplifications are difficult.

Results: Based on our previous finding that Tn5 transposase can directly cut-and-tag DNA/RNA hetero-duplexes, we present SHERRY2, a specifically optimized protocol for scRNA-seq without second-strand cDNA synthesis.

Improving Accuracy in Mass Spectrometry-Based Mass Determination of Intact Heterogeneous Protein Utilizing the Universal Benefits of Charge Reduction and Alternative Gas-Phase Reactions.

In mass analysis of proteins, mass spectrometry directly measures the mass to charge ratios of ionized proteins and promises higher accuracy than that of indirect approaches measuring other physicochemical properties, provided that the charge states of detected ions are determined. Accurate mass determination of heterogeneously glycosylated proteins is often hindered by unreliable charge determination due to the insufficient resolution of signals from different charge states and inconsistency among mass profiles of ions in individual charge states. Limited charge reduction of a subpopulation of proteoforms using electron transfer/capture reactions (ETnoD/ETnoD) solves this problem by narrowing the mass distribution of examined proteoforms and preserving the mass profile of the precursor charge state in the reduced charge states.

Evaluation of the reliability of MS1-based approach to profile naturally occurring peptides with clinical relevance in urine samples.

Rationale: The profiling of natural urinary peptides is a valuable indicator of kidney condition. While front-end separation limits the speed of peptidomic profiling, MS1-based results suffer from limited peptide coverage and specificity. Clinical studies on chronic kidney disease require an effective strategy to balance the trade-off between identification depth and throughput.