Toolkit Items

Single-cell RNA sequencing (scRNA-seq) is a transcriptomic assay method that measures RNA molecules in individual cells by sequencing-based transcript detection. In the cited application, it detected FLiCRE transcripts within the endogenous transcriptome, enabling simultaneous readout of cell type and calcium activation history.

AsLOV2 is the light-oxygen-voltage 2 photosensory domain from Avena sativa phototropin 1 used as a blue-light-responsive actuator in engineered fusion proteins. Blue-light activation drives allosteric conformational extension involving sequential unfolding of the N-terminal A'α helix and the C-terminal Jα helix, enabling conformational uncaging and related optogenetic control.

This review examines recent advancements in nanoparticle( s) (NPs) delivery systems, with a focus on ... lipid nanoparticles (LNPs)... We discussed various NP platforms and their applications, such as ... dry powder formulations of mRNA-loaded LNPs for pulmonary delivery, and LNP-mediated siRNA delivery for respiratory infections.

RNA sequencing (RNA-seq) is a transcriptomic assay method that quantifies gene-expression changes by sequencing RNA-derived libraries. In the cited study, it was used on adult rat amygdala tissue to detect subtle expression changes associated with development, cellular function, and nervous system disease after gestational high-THC cannabis smoke exposure.

The LOV2 domain of Avena sativa phototropin 1 is a blue-light-responsive protein domain that uses an FMN-dependent photocycle to reversibly switch between dark and lit states through formation and decay of a flavin-cysteinyl adduct. It has been repurposed as a modular photoswitch to control nuclear import/export motif exposure and to generate light-dependent inhibitory peptides.

The CRISPR/Cas9 system is a multi-component genome engineering platform derived from a bacterial defense system that uses Cas9 and guide RNA to manipulate genomic loci in living cells. It has been widely adopted for mutagenesis and genome research, with reported applications spanning basic biology, biotechnology, agriculture, medicine, epigenetic perturbation, and disease models.

CRISPR/Cas9 is a bacterial type II genome editing system repurposed as a programmable nuclease for target DNA cleavage and site-specific genome modification. The supplied evidence states that it was engineered for gene editing in mammalian cells by 2013 and is used to interrupt gene expression through cleavage of target DNA.

iLID/SspB is a blue-light-inducible heterodimerization system built from an engineered iLID module and the SspB binding partner. It is used to reversibly recruit proteins in cells for control of localization and signaling, including membrane recruitment, neurotrophin receptor construction, microtubule plus-end targeting, and perturbation of small GTPase pathways.

Three-dimensional (3D) bioprinting is a rapidly evolving technology that uses complementary biomaterials to emulate native extracellular matrices, enabling the generation of finely patterned, multicellular tissue architectures.

Many draw inspiration from widely successful fluorescence-based techniques such as stimulated emission depletion (STED) microscopy, photoactivated localization microscopy (PALM), and stochastic optical reconstruction microscopy (STORM).

The supplied source summary states that the review explicitly covers SIM and includes linear and nonlinear SIM.

FRASE-bot is an in silico fragment-based hit-finding method for drug discovery against unconventional therapeutic targets. It mines thousands of 3D protein-ligand complex structures to build a fragment-in-structural-environment database, matches target protein environments to that database, and uses machine learning to prioritize seeded fragments as candidate binders.

Emerging synthetic biology tools, such as CRISPR-based transcriptional control, high-throughput screening, and machine learning-assisted promoter design, are enabling the creation of tunable, orthogonal promoters suited for complex multigene expression.

incorporated into second-generation chimeric antigen receptor (CAR) constructs that were termed sherpabody-guided CARs (SbCAR)

Spatial transcriptomics is a transcriptomic assay method identified in the supplied review as a recent methodological advance. In that evidence, it is presented as part of a broader technology set that enables easier and more accurate visualization of cell behavior and qualitative and quantitative analysis of cell-cell interactions.

Chrimson is a red light-activated channelrhodopsin with a reported crystal structure. It provides red-shifted optogenetic excitation and has been used with Chronos to support two-color activation of independent neural populations in mouse brain slice without detectable cross-talk.

We focus primarily on three techniques, optogenetic manipulation, fiber photometry and microendoscopic imaging

OptoSTIM1 is an optogenetic protein tool engineered by combining the STIM1 SOAR region with a plant photoreceptor LOV2 domain. It manipulates intracellular Ca2+ levels by light-dependent activation of endogenous Ca2+-selective CRAC channels.

Phage display is an assay and selection method used during engineering workflows for light-responsive protein tools. In the cited context, it is applied alongside computational protein design and high-throughput binding assays in development of LOV2-based optogenetic systems such as improved light-induced dimers.

Many draw inspiration from widely successful fluorescence-based techniques such as stimulated emission depletion (STED) microscopy, photoactivated localization microscopy (PALM), and stochastic optical reconstruction microscopy (STORM).

fenestrations were only discernible with EM, but now they can be visualized ... in fixed cells using single molecule localization microscopy (SMLM) techniques such as direct stochastic optical reconstruction microscopy

Here we develop a proteomic kinase activity sensor technique (ProKAS) for the analysis of kinase signaling using mass spectrometry.

our optimized prime editing strategy provides a highly efficient and versatile framework for genome engineering in vitro

ProKAS is based on a tandem array of peptide sensors with amino acid barcodes that allow multiplexed analysis for spatial, kinetic, and screening applications. We engineered a ProKAS module to simultaneously monitor the activities of the DNA damage response kinases ATR, ATM, and CHK1 in response to genotoxic drugs.

The allelic series of Cry mutants is a panel of 22 cryptochrome variants, each carrying a single amino acid substitution identified by random mutagenesis and a cell-based screen. The series was used to generate diverse deficiencies across CRY functions and to identify residues involved in protein-protein interaction and CRY2-specific repression.

Amine functionalization of the membrane pore with a hydrogel exhibited >70 % retention of 20 nm negatively charged particles even when λ>40 and 80 % retention of DNA and protein when λ>160.

The CpbHLH gene family is the set of basic helix-loop-helix transcription factor genes identified in Chimonanthus praecox. A genome-wide study reported 131 CpbHLH genes distributed across 11 chromosomes and characterized their expression across tissues and flower developmental stages.

Here, we developed a genetically encoded biosensor, cdiGEBS, based on the transcriptional activity of the c-di-GMP-responsive transcription factor MrkH.

The development of the cell-free system (CFS) is transforming the manufacturing landscape of biomolecules with therapeutic value by providing a flexible and convenient alternative to cell-based expression systems.

This chapter explores the principles, platforms, and applications of CFS-based HTS... Altogether, CFS-based HTS offers a flexible, rapid, and accessible approach for next-generation biomolecular screening and therapeutic development.

cfDNA fragmentomics evaluation is an assay method that analyzes plasma cell-free DNA fragment length distributions and fragment end motifs to identify signatures associated with active gene expression. In a 2024 study, integrating short-fragment frequency with end-motif information improved enrichment for highly expressed genes in plasma samples from lung cancer patients and healthy individuals.

We report a novel dual-receptor lateral flow biosensor (LFB) for the rapid, sensitive, and visual detection of MCF-7 breast cancer cells as a model for circulating tumor cells (CTCs).

FRASE, also described as FRASE-bot, is a computational fragment-based ligand discovery method that mines 3D ligand–protein complex structures to build a database of fragments in structural environments. It screens this database against a target protein, seeds the target structure with relevant ligand fragments, and uses a neural network to prioritize fragments with the highest likelihood of being native binders.

EPR imaging further demonstrated superior diffusion of liposomal CBN through a gelatin-based semi-solid model compared to the control solution. While the current model does not replicate skin architecture, it provides a cost-effective and reproducible platform for early-stage screening of formulation mobility.

In this study, we screened a panel of 11 CjCas9 orthologous using a GFP activation assay and identified seven active nucleases.

GntR is a gluconate-responsive transcriptional repressor from Escherichia coli that has been repurposed as a protein domain for synthetic gene-control switches. Reported designs use GntR to construct gluconate-regulated transcriptional systems in mammalian cells, including rewired OFF/ON transcriptional architectures and a split transcriptional activator.

H3K36me3 cell-free chromatin immunoprecipitation sequencing (cfChIP-seq) is a plasma-based assay that establishes a personal gene expression profile from cell-free chromatin. In the cited study context, it functions as a reference enrichment assay for active genes in liquid biopsy samples.

H3K36me3 cfChIP followed by droplet digital PCR is a cell-free chromatin immunoprecipitation assay that enriches plasma cfDNA associated with the transcription-linked histone mark H3K36me3 and then quantifies specific alleles by ddPCR. In a 2021 NSCLC study, it detected greater enrichment of EGFR-L858R fragments than EGFR wild-type fragments, providing proof of principle for identifying tumor-specific transcriptional activity of mutated alleles.

we developed a simple, rapid, and efficient system based on hairy root transformation to evaluate somatic genome editing efficiency in plants

The high-throughput online monitoring system with an LED array is an assay platform for screening light-controlled gene expression conditions by individually illuminating each well in a multiwell format. In the cited yeast study, it was used with photocaged Cu2+ to regulate the Cu2+-inducible pCUP1 promoter from Saccharomyces cerevisiae and monitor eYFP expression.

High-throughput screening is an assay method cited in microbial biotechnology literature as part of the CRISPR/Cas toolbox for evaluating variants generated by multiplexed engineering. In the supplied evidence, it is presented as a screening approach associated with CRISPR/Cas-based metabolic engineering and with development of new dynamic systems.

In this work, we present "inkube", an incubation system that has been combined with an electrophysiology setup and a fully automatic perfusion system.

LC-MS analysis of fittest binders is an assay method used with small combinatorial libraries of self-assembled proteomimetics (SAPs) to identify enriched target binders after affinity selection by liquid chromatography–mass spectrometry. In the cited SAP study, this workflow was applied in the context of target-directed selection from self-assembled PNA-peptide conjugate libraries.

we developed a biosensor based on turn-on fluorescent protein, named LV-EcpG

A functional lentiviral vector, LV_EF1α_GBA_Opt, was generated at a titer of 7.88 × 108 LV particles/mL as determined by qPCR.

Screening of antioxidant properties of compounds needs appropriate methods including metal chelating assay.

The review integrates data from in vitro, in silico, and clinical studies, including both classical detection strategies and emerging technologies such as clustered regularly interspaced short palindromic repeats (CRISPR)-based modulation, biosensors, and microfluidics.

Pseudovirus technology, which uses single-round infectious viral particles lacking replication competence, has thus gained prominence as a safe and versatile tool for antiviral research.

This directed protein evolution generates several gain-of-function OsTIR1 variants, including S210A, that significantly enhance the overall degron efficiency.

Among them, paper-based biosensors have emerged as a promising platform due to their low fabrication cost, simplicity, biodegradability, and compatibility with point-of-care (POC) testing.