Electrophoretic mobility shift assay (EMSA, also called gel retardation assay or gel shift assay) is an in vitro method to detect the interaction between proteins and nucleotides. It is originally used to detect transcription factors, and is now further developed into investigating DNA-protein interactions, RNA-protein interactions, and even DNA-RNA interactions. Profacgen provides professional EMSA service to identify potential DNA or RNA binding proteins with high efficiency and accuracy.
The principle of EMSA is based on the electrophoretic mobility discrepancy between biomolecules with different molecular masses. Nucleotides move slower during electrophoresis when bound with proteins, thereby giving a lagged band in the final detection. EMSA can be used to qualify and quantify proteins that specifically bind to given nucleotides. We can evaluate the binding affinity according to the quantity of the lagged band, and even calculate the associate and dissociate constant. EMSA is often performed concurrently with DNase I footprinting and primer extension experiments when studying transcription initiation, DNA replication, DNA repair or RNA processing and maturation.
Schematic Diagram of EMSA
Profacgen has a mature EMSA experiment platform and extensive experience in nucleotide-protein binding analysis. We perform strict examination on both nucleotide and protein to ensure they are qualified for EMSA before the initiation of each project. We can use radioactive isotope and also other labeling agents such as biotin and FAM to label nucleotides. Unlabeled nucleotides are used as specific competitors to eliminate non-specific nucleotide-protein interactions. We also have advanced instrument for signal detection ensuring high sensitive and high signal-to-noise ratio results.
Protein sample: We normally require the purity of the protein sample to be >90%, and concentration >0.3 mg/ml.
Cell/tissue samples: Cell sample should include >107 cells; tissue sample should be >100 mg.
Nucleotides: Nucleotides used as probes should be sub-cloned into plasmids and verified by DNA sequencing; or you can simply provide the nucleotide information (gene name, source, GenBank accession number, sequence, etc.)
Service work flow:
Please contact us for our professional EMSA service. Our experienced scientists will provide you with tailered tailored experimental scheme for your specific requirements.
References from our customer:
Sirisena, Nirmala D., Nilakshi Samaranayake, and Vajira HW Dissanayake. "Electrophoretic mobility shift assays implicate XRCC2: rs3218550C> T as a potential low-penetrant susceptibility allele for sporadic breast cancer." BMC research notes 12.1 (2019): 476.
XRCC2 is known to play an essential role in homologous recombination repair of DNA double-strand breaks. It is plausible that this variant may be exerting regulatory effects on XRCC2 gene expression leading to altered DNA repair capacity. Further functional studies are warranted to validate this finding.EMSA was performed by Profacgen.