A

Aggregates

Non-functional clumps of proteins that resemble condensates but lack liquid-like properties. Current TRAPS experiments show aggregation that must be resolved.

Auxotrophic Markers

Genes used in yeast or other organisms that allow selection of transformed cells by complementation of a nutritional deficiency, e.g., URA3 or LEU2.

B

Bead Beating

A mechanical cell disruption method using small silica beads and vigorous shaking to physically break cells apart.

Buffer

Solutions used to maintain pH and ionic strength in experiments.

C

Cas13

An RNA-targeting CRISPR protein. In TRAPS, a catalytically inactive form (dCas13) binds RNA without cutting it, acting as a programmable sensor.

Crude Extract

The unpurified mixture obtained after lysing cells; contains proteins and other cell components.

Condensates (Biomolecular Condensates)

Membrane-less compartments formed by phase separation of proteins and/or RNAs. In TRAPS, condensates act as organisational hubs for RNA detection by clustering Cas13 molecules.

D

dCas13

A mutated version of Cas13 that binds RNA without cleavage activity. Guides condensate formation to specific RNA sequences.

Densitometric Analysis

A quantitative method used to measure the intensity of bands or spots (such as proteins or nucleic acids) on gels or blots. The darkness or brightness of each band, captured in an image, is analyzed using software to estimate relative amounts of the target molecule.

E

E9–Im2 Scaffold

A synthetic protein interaction system adapted from Heidenreich et al. (2020). Provides the structural basis for TRAPS condensate formation.

Exponential Phase

The period of rapid cell growth and division when cells are healthiest and actively multiplying.

F

Fluorophore (GFP/mCherry)

Fluorescent proteins used to tag TRAPS scaffolds and RNA targets for visualization.

FRAP (Fluorescence Recovery After Photobleaching)

A microscopy technique used to study the mobility of fluorescently labeled molecules in living cells. A defined region is bleached with intense light, and the recovery of fluorescence is monitored as unbleached molecules move back into the area. This reveals diffusion rates and binding dynamics.

G

Galactose Promoter (GAL1)

An inducible promoter in yeast used to control expression of mCherry RNA for TRAPS validation experiments.

Gateway Cloning

Gateway cloning is a recombination-based DNA cloning method that avoids restriction enzymes and ligases. It uses site-specific recombination from bacteriophage λ, where DNA fragments flanked by special recombination sites (att sites) are transferred between vectors. The process occurs in two steps: the BP reaction, which creates an entry clone, and the LR reaction, which transfers the insert into a destination (expression) vector.

GFP (Green Fluorescent Protein)

A green fluorescent protein tag fused to TRAPS scaffold proteins to visualize condensates.

gRNA (guide RNA)

A short RNA sequence that directs Cas13 to its RNA target. Multiple gRNAs improve binding and condensate formation.

H

Histidine Tag

A small amino acid tag used for protein purification and detection.

I

Im2

Interaction partner in the E9–Im2 scaffold system that allows condensate assembly.

Immunoblotting (Western Blotting)

A method to detect specific proteins separated by SDS-PAGE, using antibodies on a membrane.

Immunostaining

A technique using antibodies to detect specific proteins in cells or tissues, often visualized by fluorescence or color.

L

LLPS (Liquid–Liquid Phase Separation)

A specific type of phase separation in cells where proteins and RNAs form dynamic, liquid-like condensates. These membraneless organelles concentrate biomolecules to regulate biochemical reactions and can assemble or dissolve rapidly.

Lysis Buffer

A chemical solution used to break open cells and release their contents (proteins, DNA, etc.) for analysis.

M

mCherry

A red fluorescent protein used as a proof-of-concept RNA target in yeast.

P

Phase Separation

A process where a homogeneous mixture separates into distinct phases with different compositions. In biology, it often refers to the segregation of biomolecules into concentrated droplets or compartments without membranes.

Pulldown Assay

A biochemical method to test protein–protein or protein–nucleic acid interactions. One molecule (the “bait”) is immobilized on beads, and potential binding partners (the “prey”) are captured from a mixture. After washing away non-specific binders, the interacting partners are analyzed, often by SDS-PAGE or mass spectrometry.

Pumby

A protein derived from the Pumilio family that binds RNA sequences with high specificity. Can be used as a modular RNA-binding domain in synthetic biology.

O

OD₆₀₀ (Optical Density at 600 nm)

A spectrophotometric measure of cell density in a culture; higher OD₆₀₀ means more cells.

P

Phase Separation

A process where a homogeneous mixture separates into distinct phases with different compositions. In biology, it often refers to the segregation of biomolecules into concentrated droplets or compartments without membranes.

Pulldown Assay

A biochemical method to test protein–protein or protein–nucleic acid interactions. One molecule (the “bait”) is immobilized on beads, and potential binding partners (the “prey”) are captured from a mixture. After washing away non-specific binders, the interacting partners are analyzed, often by SDS-PAGE or mass spectrometry.

R

rcf (Relative Centrifugal Force)

The force applied to samples in a centrifuge, expressed in multiples of gravity (× g); determines how strongly particles are spun down.

RNA

Ribonucleic acid, a molecule that carries genetic information and can act as a template for protein synthesis. In TRAPS, target RNA sequences are detected by dCas13.

rpm (Revolutions per Minute)

The speed of centrifuge rotation; must often be converted to rcf depending on rotor size.

S

S. cerevisiae (W303)

The yeast model organism used to test TRAPS. Chosen for its ease of genetic manipulation.

SD -TRP -URA Medium

A type of synthetic defined (SD) yeast growth medium lacking the amino acid tryptophan (TRP) and nucleotide uracil (URA); used to select for yeast strains carrying plasmids that supply these nutrients.

SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis)

A technique to separate proteins by size using an electric field.

Signal-to-Noise Ratio (SNR)

A measure of how strong a desired signal is compared to background noise. In imaging or data analysis, a higher SNR means clearer, more reliable results, while a low SNR makes it difficult to distinguish true signals from random fluctuations.

Supernatant (S)

The liquid layer remaining after centrifugation, containing soluble components.

T

Tag

Short peptide sequence or protein added to a target protein to aid in detection or purification (e.g. myc tag, FLAG tag).

Tetramerization Domain

A protein domain fused to GFP and E9 in TRAPS, forming fluorescent tetrameric scaffolds that can be linked together by RNA-bound Cas13.

Total Fraction (T)

A sample representing the entire cell lysate before centrifugation or separation.

TRAPS (Targeted RNA Activated Phase Separation)

A modular system for detecting RNA in living cells by coupling dCas13 to phase-separating scaffolds that form fluorescent condensates when the target RNA is present.

W

Western Blot

A laboratory method used to detect specific proteins in a sample using antibodies. Useful for verifying expression of TRAPS scaffolds or fusion proteins.

Y

YPD Medium/YEPD Medium

Yeast Extract Peptone Dextrose; a complete medium for yeast growth containing yeast extract, peptone, and dextrose (glucose); supports fast, non-selective growth.