Toolkit Items

A NIMPLY B gates are compressed two-input mixed-phenotype transcriptional logic operations reported in a 2023 ACS Synthetic Biology study on performance prediction of fundamental transcriptional programs. The study indicates that their behavior can be modeled and predicted from experimentally characterized single-input logical operations and associated metrology.

Accelerated MD simulation is an in silico computational method reported for elucidating the photoactivation mechanism of the AsLOV2 light-responsive domain. The available evidence supports its use as a mechanistic analysis approach for a protein photosensor rather than as a deployable biological reagent.

All-atom replica exchange discrete molecular dynamics is a computational docking method used to generate structural models of calcium and integrin binding protein 1 (CIB1) bound to α-integrin cytoplasmic tails. In the cited CIB1 study, it predicted that multiple α-integrin tails engage the same hydrophobic binding pocket on CIB1.

AlphaFold3 is a computational structure-prediction method used in the cited study to model the MagMboI–DNA complex. In that work, it was applied to infer interactions with the 5'-GATC-3' recognition sequence and to guide optimization of the photoactivatable endonuclease variant MagMboI-plus for top-down genome engineering.

The Anderson-Darling test is a nonparametric computational method applied in genome-wide association studies of complex quantitative traits. In an enlarged maize association panel, it identified loci across 17 agronomic traits, including both previously known loci and additional candidate loci detected only by this test.

B NIMPLY A gates are compressed mixed-phenotype two-input transcriptional logic operations reported in a 2023 ACS Synthetic Biology study on performance prediction of fundamental transcriptional programs. In that context, their behavior was modeled and predicted from single-input data and metrology rather than described as a separately detailed molecular construct.

This Bayesian computational approach is a data-analysis method developed to improve prediction of split protein behavior by contextualizing errors inherent to experimental procedures. In the cited study, it was applied to pooled, sequencing-based screening data from split Cre recombinase constructs generated with optogenetic dimers, enabling comprehensive analysis of split sites across the protein.

The Bayesian optimization framework is a computational method built from high-throughput Lustro measurements to guide control of blue light-sensitive optogenetic systems. It uses data-driven learning, uncertainty quantification, and experimental design to identify light induction conditions for multiplexed regulation in Saccharomyces cerevisiae.

The binding equilibrium model is a computational modeling approach used to quantitatively describe how proteins partition into engineered synthetic condensates. In the reported synthetic membraneless organelle framework, it supports prediction of condensate composition based on affinity-dependent recruitment.

This computation method is a bioinformatic analysis applied to protein components of the Saccharomyces cerevisiae ribosome assembly pathway. In the cited study, it compared ribosome biogenesis factors with ribosomal proteins and found that biogenesis factors contain significantly more predicted trans interacting regions.

Blue-light controlled gene modules are optogenetic expression constructs used with a portable smart blue-light controlled device to regulate local cytokine expression in the tumor microenvironment. In the cited 2024 MED study, this strategy was associated with enhanced activation of tumor-infiltrating immune cells and remodeling of an immunosuppressive tumor milieu.

BROAD is a computational protein design method that combines Rosetta-based structure modeling, machine learning, and integer linear programming to improve design search beyond Rosetta sampling alone. It was demonstrated in antibody design to increase the predicted HIV neutralization breadth of VRC23 across a panel of 180 divergent viral strains.

The C-terminal extension is an exceptionally long structural feature characteristic of plant cryptochromes, which are blue-light receptors containing flavin adenine dinucleotide within a photolyase homology region. Current mechanistic models place this extension downstream of light-induced flavin reduction, with signaling culminating in dissociation of the C-terminal extension.

Caspase-2 active-site mutants are engineered variants of human Caspase-2 designed in silico to broaden substrate recognition at the substrate N-terminal amino acid position. In vitro experiments confirmed that two proposed mutants showed enhanced promiscuity, including increased recognition of branched amino acids relative to unmutated Caspase-2.

The catalytic/ATP-binding (CA) domain is the kinase catalytic module of the light-regulated histidine kinase EL346. In the reported structural model, its activity is regulated indirectly by LOV-domain photoactivation through changes in the DHpL/CA interface that release the CA domain from an inhibited dark-state conformation.

Chemogenetically driven repositioning of lysosomes is an experimental perturbation used to causally alter lysosome localization and test how lysosome positioning regulates endoplasmic reticulum remodeling. In the cited 2020 Science Advances study, chemo- and optogenetically driven lysosome repositioning was used to validate a causal link between lysosome movement and ER network organization.

ChETA is a light-responsive protein domain considered here only in a theoretical co-expression strategy with step-function opsins (SFOs). In that context, modeling predicted that ChETA-SFO co-expression in hippocampal neurons would stabilize photocurrent and reduce spike failure associated with photocurrent desensitization.

ChR2(C128S) is a step-function channelrhodopsin variant identified as one of several step-function opsins considered in a 2022 theoretical optogenetics study. In that study, it is modeled in co-expression with the fast channelrhodopsin ChETA to support sustained neuronal switching and improved spiking performance.

CIB1-targeting decoy peptides are computationally modeled peptide variants intended to bind calcium and integrin-binding protein 1 (CIB1) and inhibit its function. A 2023 in silico study reported that top-ranked second-generation mutant peptides had greater predicted inhibitory potential than the reference peptide UNC10245092.

The Cib1−/− mouse model is a genetic loss-of-function system in which calcium- and integrin-binding protein 1 (CIB1) is deleted to study its role in megakaryocyte and platelet biology. In the cited Blood 2011 study, Cib1 deficiency altered megakaryocyte adhesion, migration, and proplatelet formation and attenuated platelet recovery after depletion.

Clathrin endocytosis is a host-cell internalization pathway described in a review as one route used by Kaposi’s sarcoma-associated herpesvirus (KSHV) to enter fibroblast infection models. In this context, it serves as a cellular uptake mechanism associated with viral entry and subsequent trafficking events linked to host signaling pathways.

Compressed AND gates are two-input transcriptional logic operations within a framework of fundamental compressed logical programs. In the cited ACS Synthetic Biology study, their behavior was modeled and predicted from experimentally characterized single-input gate data and associated metrology rather than reported as a separately engineered standalone construct.

Compressed NOR gates are predicted two-input transcriptional logic operations within a framework of fundamental compressed logical programs. In the cited ACS Synthetic Biology study, their behavior was modeled from experimentally characterized single-input logical elements and associated metrology rather than reported as directly constructed standalone gates.

This computational design strategy combines in silico design with biophysical experiments to improve the response kinetics of protein conformational switches. In the cited 2018 Nature Communications study, it was applied to an engineered protein-based Ca2+ sensor and rationally accelerated its response into the range of fast physiological Ca2+ fluctuations.

Computational methods for LOV-based optogenetic tool development are design-enabling approaches used in the ongoing development of Light-Oxygen-Voltage domain-based optogenetic systems. The cited evidence supports a role for computational methods as one of several factors advancing LOV-based tools for light-controlled biological regulation.

Computational modeling was used to analyze how promoters decode light-driven transcription factor nuclear translocation dynamics. In the cited work, the modeling identified promoter kinetic regimes that enable efficient expression under short light pulses and proposed a multi-stage, thresholded activation scheme to explain opposite promoter-response phenotypes.

Computational protein design is an engineering methodology described in a 2018 review as a next-generation tool for expanding synthetic biology applications. The supplied evidence frames it as a design approach used alongside phage display and high-throughput binding assays rather than as a single molecular reagent.

Conditional gene mutation is a genetic engineering method for creating gene alterations that are activated in spatially restricted and/or temporally restricted ways. In mice, these conditional mutations enable controlled genetic perturbation for experimental studies and disease modeling.

Conventional replica exchange molecular dynamics (REMD) is a molecular simulation method used for statistical sampling of biomolecular conformational ensembles. In the cited evidence, it functions as the benchmark comparator against two coarse kMC-based replica exchange methods.

CRISPR/Cas9 is a genome editing technique used in the cited study to generate Cib1 and Cib2 knockout mice. In this evidence set, its demonstrated function is targeted gene disruption for mouse model production.

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.

CRY-BARs are a family of light-gated optogenetic tools that contain an I-BAR domain and are designed to remodel membrane architecture. They are intended to provide spatial and temporal control over cellular processes involving membrane protrusion formation.

CRY1 knockout cells are human U-2 OS osteosarcoma cell models in which the CRY1 locus was disrupted using duplex CRISPR/Cas9 editing. The reported approach uses two guide RNAs targeting intronic regions flanking a CRY exon to remove whole exons and generate a human circadian clock knockout model.

CRY1/CRY2 double knockout cells are human U-2 OS osteosarcoma cell models in which both cryptochrome genes were disrupted using duplex CRISPR/Cas9. The tool was generated by deleting whole exons with paired guide RNAs targeting exon-flanking intronic regions, producing a double-knockout circadian phenotype reported as arrhythmic.

CRY2 knockout cells are human U-2 OS osteosarcoma cell models in which the CRY2 gene was disrupted using duplex CRISPR/Cas9. They were generated by deleting whole CRY exons with two guide RNAs targeting exon-flanking intronic regions and display long-period circadian phenotypes consistent with CRY loss.

Designed decoy peptides targeting CIB1 are in silico engineered variants of the reference peptide UNC10245092 generated by residue scan methodology to bind CIB1. In the reported computational study, top candidates were prioritized by predicted binding free energy, evaluated by AMBER molecular dynamics simulations, and were predicted to interfere with RAF–CIB1 binding; mutant 2 showed stronger predicted interactions with CIB1 than the reference peptide.

The Diaphanous Autoregulatory Domain from mDia1, in this tool context, is an optogenetic fusion between the Avena sativa Phototropin1 LOV2 domain and an isolated mDia1 DAD. Blue light uncages the DAD, enabling rapid activation of endogenous diaphanous-related formins and acute actin cytoskeletal remodeling.

DiLiCre is an optimized photoactivatable Cre recombinase system described as a doxycycline- and light-inducible Cre recombinase mouse model for optogenetic genome editing. It enables inducible Cre-mediated genome recombination in vivo, including light-induced mutagenesis and positional cell tracing by intravital microscopy.

Duplex CRISPR/Cas9 technology is a genome-editing method that uses two guide RNAs to target intronic sequences flanking an exon, enabling excision of the intervening exon by Cas9-mediated cleavage. In human U-2 OS osteosarcoma cells, it was applied to generate CRY1 knockout, CRY2 knockout, and CRY1/CRY2 double knockout cell models.

Dynamic multiplexing is a computational design principle for synthetic gene networks that encodes and decodes time-varying inputs into distinct gene expression states. In the cited 2021 study, it increased information transmission from signal to gene expression and enabled dynamic signal decoding using engineered regulators with different response kinetics.

The elastic network model (ENM) is a computational method for analyzing protein structural dynamics from an elastic network representation. In the cited PAS domain superfamily study, ENM was used to quantify residue fluctuations and vibrational patterns and to relate these dynamic features to sequence conservation, structural features, and biological function.

Electron-electron double resonance (ELDOR) spectroscopy is a structural assay method that, when combined with site-directed spin labelling, was used to chart light-induced structural transitions in the engineered LOV histidine kinase YF1. In the cited study, it provided pairwise distance information used to model blue-light-driven quaternary rearrangements in a signaling photoreceptor.

Engineered BUFFER (repressor) and engineered NOT (antirepressor) logical operations are single-input transcriptional logic elements developed as network-capable components for synthetic gene programs. They were experimentally characterized and used as foundational parts for predictive modeling of more complex compressed transcriptional logic circuits.

This tool is an engineered protein-based Ca2+ sensor that switches through mutually exclusive folding of two alternate frames. Computational design and biophysical experiments were used to improve its conformational switching kinetics, producing Ca2+-responsive behavior on the timescale of fast physiological Ca2+ fluctuations.

Exendin-4 fused with an anti-neonatal Fc receptor affibody is a therapeutic fusion payload used in the ROEN red light-controlled probiotic system. In the cited murine study, this construct was released under optogenetic control in the gut to mediate gut-brain axis regulation and was evaluated in a Parkinson's disease mouse model after oral administration.

Fernando's model is a computational model of a synthetic molecular circuit designed to mimic Hebbian learning in a neural network architecture. It is described as one of the earliest models in this area to use Hill equation-based regulatory modeling, and computational analysis indicated that a reinforcement effect can be obtained with appropriate parameter choices.

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.

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.

Free-energy calculations are an in silico prediction method used in the rational design of human Caspase-2 mutants. In the cited study, they were applied alongside sequence and structural comparisons of Caspase-2 and Caspase-3 to predict effects of active-site mutations on substrate recognition and to support engineering of broader amino-acid acceptance.

The GelMA-Macrophages-LED system is a composite delivery harness composed of a light-crosslinked GelMA hydrogel, gene-modulated macrophages, and a wireless LED device. In the cited study, it was used for in situ light regulation of cardiac inflammation in murine lipopolysaccharide-induced sepsis models, with macrophage photoactivation linked to suppression of inflammatory cytokine production.