Experimental Details and the TRANSPATH Quality Matrix

Details about the materials and methods used to determine interactions between and among TRANSPATH molecules are curated, when the specific information is provided in the literature. Descriptions of materials and methods curated in TRANSPATH are provided in the tables below.


Code Source Description
A1 Wild type These terms describe a whole organism or its parts. The terms can refer to a wild-type organism, a naturally-occurring mutant organism, or a genetically manipulated organism.
A2 Naturally-occurring chromosomal mutants
A3 Knock out
A4 Transgenic organism
A5 Organism with induced but not defined mutation(s)
B1 Embryonic cells These terms describe primary cultures of cells, possibly transfected. The terms do not describe permanent cell lines.
B2 Stem cells -> pluripotent
B3 Primary cells
B4 Recombinant cells/tissue/organisms (transfected); not permanent cell lines
C1 Permanent tissue culture cells Permanent cell lines (permanent cultures of cells).
D1 Permanently or transiently transfected or infected cells (tissue culture cells) Vertebrate or insect cells expressing recombinant protein.
D2 Insect or Xenopus cells producing recombinant protein (for example, Baculovirus vectors in insect cells or transfected/microinjected Xenopus oocytes)
E1 Yeast cells producing recombinant protein Recombinant protein expressed in yeast cells.
F1 Bacteria producing recombinant protein Recombinant proteins expressed in bacterial cells or synthesized in vitro.
F2 in vitro translation, synthetic peptides
X1 Unspecified cell type producing recombinant protein --
X2 Unspecified material --

Additional information about TRANSPATH materials, including examples, can be found here.


Code Method Description/examples
M1 Yeast two hybrid assay --
M2 Screening of expression libraries with other (labeled) proteins Phage display, eukaryotic cDNA library with EXIox vectors
M3 (Co-)localization of proteins in cells and tissues by microscopical analysis Immunofluorescence, in situ detection, electron microscopy
M4 Affinity precipitation in solution/batch (with sepharose or magneto beads, for example) Immunoprecipitation, pull-down assay
M5 in vitro protein-protein binding assay (solid phase) Enzyme Linked Immuno Sorbent Assay (ELISA)
M6 Surface plasmon resonance Detection and quantification of ligands
M7 Far Western, Gel overlay, or protein-lipid overlay assay Gel overlay: detection of binding partners on blotted proteins after transfer from a gel
M8 Affinity chromatography with matrices Proteins coupled to sepharose or agarose matrices, for example
M9 Peptide spot Search for binding partners of defined peptide motifs on a membrane
M10 Cross linking Detection by SDS-Page and Western blot
M11 Electrophoretic mobility shift assay (EMSA) --
M12 Binding assay Binding of labeled ligand to receptor-expressing cells, quantitative evaluation, for example Scatchard analysis
M13 Modification of proteins in living cellss Protein phosphorylation and -dephosphorylation, ubiquitination, farnesylation; detection of the modified proteins from cell extracts
M14 Subcellular extraction of proteins Fractionation for localization of proteins
M15 Dot spots Spotted proteins detected with specific antibodies
M16 in vitromodification of a substrate Phosphorylation or dephosphorylation (kinase assay), ubiquitination
M17 Reporter gene assay Luciferase, beta-galactosidase assay
M18 Western blot, SDS page --
M19 mRNA detection by Northern blot Indirect evidence for protein interaction
M20 Localization of mRNA in tissues by e.g. FISH, ISH (p32, H3) Indirect evidence for protein interaction
M21 Detection of mRNA by RT-PCR Reverse transcriptase, thereafter PCR; quantitative mRNA detection; indirect evidence for protein interaction
M22 Mammalian two (or tri) -hybrid assay --
M23 Co-sedimentation and -purification by chemico-physical methods Biochemical purification of stable complexes by e.g analytical ultracentrifugation, size-exclusion chromatography (SEC)
M24 Protein cleavage and specific degradation Documented by detection of labeled protein fragments, immunoblots, SDS-PAGE, HPLC, mass spectral analysis
M25 Interaction measurement by chemico-physical methods Crystal structure analysis of complexes; spectrofluorometer fluorescence resonance energy transfer (FRET); intensity and/or wavelength of fluorescence of a given interacting compound changes, when another molecule binds to it; quantitative; usually done in vitro; also NMR: looking at changes in nuclear spin, usually done with peptide fragments and in unphysiological solutions, isothermal titration calorimetry (ITC)
M26 mRNA expression profile analysis Microarray data
M27 Transmembrane potential measurements Current-voltage experiments
M28 Predicted by binding-site sequence --
M29 Co-localization --
M30 in vitro binding --
M31 in vitro reconstitution of activity --
M32 Flow cytometry --
M33 Ligand binding --
M34 Two hybrid Cell system not specified
M35 Enzyme activity measurement --
M36 DNA footprinting/interference --
Chromatin immunoprecipitation --
M38 Filter binding assay --
M39 One hybrid Cell system not specified
M40 Three hybrid Cell system not specified
M41 Osmium tetroxide modification --
M42 Southwestern blot --
M43 Bacterial two hybrid assay --
M44 Bimolecular Fluorescence Complementation (BiFC) --
M45 Immunoprecipitation of small molecule complexes with proteins e.g. receptors --
M46 Competitive binding assay of small molecule binding --
M48 Enzyme immunoassay (EIA) --
M49 RNA silencing/interference --
M50 AP-MS (affinity purification coupled with mass spectrometry) --
M51 Bioinformatic processing --
M52 XL-MS (chemical cross-linking coupled with mass spectrometry) --
M53 Proximity-Labeling MS --
M54 BioID (proximity-dependent biotin identification) --
M55 SILAC (stable isotope labeling by/with amino acids in cell culture, based on mass spectrometry) --
M56 AVA-Seq (all-versus-all sequencing based on a bacterial two-hybrid system) --
M57 PCA (Protein-Fragment Complementation Assay) --

Importantly, TRANSPATH not only extracts data about the materials and methods from the published literature; TRANSPATH also includes a quality evaluation and a reliability code for the material-method combinations. As such, each material and each method is given a particular symbol, and combinations of the symbols are given a reliability code, as shown in the TRANSPATH Quality Matrix.

Please note: Curators use only data from peer-reviewed publications. If controls are missing or are not credible in a specific paper, the data will not be included.

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