Part:BBa_K5160010

3xFLAG

Overview

The flag tag protein is a fusion peptide consisting of 8 amino acids. In protein expression and localization research experiments, the gene of interest and the flag-tag gene sequence can be linked together through genetic engineering techniques. This linkage can occur at either the C-terminus or N-terminus of the target protein, and the integrated gene is then transferred into cells for specific detection. As a commonly used protein tag in laboratories, it typically does not interact with the target protein and does not affect the function, properties, or structure of the target protein.
In project design, to purify and identify the target protein while enhancing the detection signal and improving purification efficiency, we attach three flag tag proteins to the C-terminus of the target protein to create the corresponding vector. Subsequently, we introduce the target gene into the tomato genome sequence through transient infection technology or transgenic technology. When the target gene is expressed, the flag tag protein is also expressed, which can be used for subsequent Western blot (WB) detection and co-immunoprecipitation to verify the expression of the target protein.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design

1. During the design process, considering the need to add the vacuolar signal peptide SPS-NTPP to the N-terminus of the target sweet protein, we deliberately placed the 3x Flag tag at the C-terminus of the target protein to prevent it from interfering with the SPS-NTPP vacuolar signal peptide. 2. Meanwhile, to improve the accuracy of detection, we used three flag tags. Multiple tags can increase the number of antigenic epitopes, thereby enhancing the chances of recognition by antibodies and strengthening the signal intensity of Western blot (WB) or immunoprecipitation. Additionally, multiple tags can reduce the occurrence of false-negative results in experiments. Even if one tag fails to be detected due to certain reasons, other tags may still be detectable. Furthermore, multiple tags can enhance the detection signal, improve purification efficiency, increase flexibility, reduce the risk of loss, and facilitate quantitative analysis.

Source

The flag protein is an artificially designed tag protein.