Part:BBa_K4818027
3xFLAG
The term "3xFLAG" refers to a protein tag used in molecular biology and biochemistry to facilitate the detection, purification, and characterization of proteins. It consists of three consecutive FLAG epitope tags, which are short amino acid sequences recognized by specific antibodies. Additionally, the 3xFLAG tag is often followed by an enterokinase cleavage site. Here's a breakdown of each component:
1. **FLAG Epitope Tag**: The FLAG epitope tag is a small, highly specific peptide sequence derived from a protein originally found in Escherichia coli (E. coli). The sequence that comprises the FLAG tag is DYKDDDDK. This sequence is designed to be recognized by commercially available anti-FLAG antibodies. When a protein is genetically engineered to include one or more FLAG tags, it can be easily detected and purified using these antibodies.
2. **3xFLAG**: The "3x" in 3xFLAG signifies that the FLAG epitope sequence is repeated three times in tandem within the tag. This tripling of the FLAG sequence enhances the binding affinity between the tag and anti-FLAG antibodies, making it easier to detect and purify the tagged protein.
3. **Enterokinase Cleavage Site**: Following the 3xFLAG tag, there is an enterokinase cleavage site. Enterokinase is a proteolytic enzyme that specifically recognizes and cleaves its target peptide sequence after the amino acid sequence Asp-Asp-Asp-Asp-Lys (DDDDK). This site is designed to allow for the removal of the 3xFLAG tag from the protein if needed. By treating the tagged protein with enterokinase, researchers can cleave the tag from the protein of interest, leaving it untagged.
The inclusion of the 3xFLAG tag, along with the enterokinase cleavage site, provides several advantages in protein research:
- **Detection**: The 3xFLAG tag allows for easy detection of the tagged protein using anti-FLAG antibodies in various assays, including Western blots and immunoprecipitation.
- **Purification**: Researchers can use anti-FLAG affinity chromatography to purify the protein of interest from complex mixtures. The tag facilitates the isolation of the tagged protein.
- **Flexibility**: The enterokinase cleavage site offers flexibility. Researchers can remove the tag after purification if they wish to study the untagged protein, as the presence of the tag might interfere with certain experiments or assays.
In summary, 3xFLAG is a protein tag consisting of three consecutive FLAG epitope tags followed by an enterokinase cleavage site. It is a powerful tool in molecular biology for the detection, purification, and study of proteins of interest.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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