Myc Tag, Recombinant, Western Blot Positive Control

Expression of genes in E. coli or yeast or baculovirus offers a convenient system to produce large amounts of recombinant proteins that may otherwise be difficult to isolate from natural cells and tissues. Very often antibodies to these newly identified proteins are not available to study its biochemical properties, monitor protein expression, and purification. In order to circumvent this problem, short pieces of well-defined peptides (Poly-His, Flag- epitope or c-myc epitope or HA-tag) or small proteins (bacterial GST, MBP, Thioredoxin, b-Galactosidase, VSV-Glycoprotein etc) are often cloned along with the target gene. Proteins are expressed as fusion proteins. Antibodies to these fusion-tags are already available to monitor fusion protein expression and purification. Therefore, fusion-tags serve as universal tags much like secondary antibodies. Many tags have their own characteristics. Poly-His-fusion proteins (6 x His) can bind to Nickel-Sepharose or Nickel-HRP. GST-fusion proteins can bind to glutathione-Sepharose. Therefore, a high degree of purification of fusion protein can be achieved in just one affinity purification step. Purity of fusion proteins can be followed by Tag-antibodies. Very often, fusion proteins are directly injected into animals to generate antibodies. Some fusion tags can be removed later by treatment with enzymes to generate tag-free recombinant proteins.\n\nSource:\nSynthetic 10aa peptide sequence from human myc protein (aa410-419; EQK LIS EED L). The myc-tag was expressed as N-terminal fusion protein in E. coli.\n\nMolecular Weight:\n~9.5kD\n\nApplications:\nSuitable for use in Western Blot as positive control. Not suitable for ELISA or other applications where native protein is required. Other applications not tested.\n\nRecommended Dilutions:\nWestern Blot: 10ul/lane. SDS may crystallize in cold conditions. It should redissolve by warming before taking it from the stock. It should be heated once prior to loading on gels. If the product has been stored for several weeks, then it may be preferable to add 5ul of fresh 2x sample buffer per 10ul of the Western Blot solution prior to heating and loading on gels. This preparation is not biologically active.\nOptimal dilutions to be determined by the researcher.\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20 degrees C. Aliquots are stable for 6 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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