Table of Contents
What is His-tag used for?
His Tags allow researchers to selectively extract a protein of interest from the thousands of other proteins found in either a cell or cell lysate.
How does 6 His-tag work?
Some recombinant proteins are engineered to have two hexahistidine tags. His-tag purification uses the purification technique of immobilized metal affinity chromatography, or IMAC. In this technique, transition metal ions are immobilized on a resin matrix using a chelating agent such as iminodiacetic acid.
Why is a 6 His-tag necessary for Ni-NTA purification?
The reason for using 6xHis might be the determined through trial and error, as it is the shortest stretch of polyhistidine peptide that can be purified by IMAC to satisfactory purity in high yield.
How do I get rid of His-tag?
His-tag removal from protein using TEV Protease
- Dialyze the protein against 20 mM Tris-HCl, pH 7.5.
- Determine the protein concentration.
- Combine 15 μg of protein and H2O (if necessary) to make a 45 μl total reaction volume.
- Add 5 μl of TEV Protease Reaction Buffer (10X) to make a 50 μl total reaction volume.
How do you regenerate Ni-NTA resin?
For Qiagen’s Ni-NTA, a simple regeneration protocol is:
- Wash with water.
- Remove Ni2+ ions with 50 mM EDTA.
- Wash with water.
- Clean with 0.5 M NaOH.
- Neutralise with water (this will take some time)
- Regenerate with 100 mM NiSO.
- Wash with water and then either 20% ethanol or buffer.
What is a 6x His tag?
Target Information. The 6x His tag is a synthetic oligo peptide consisting of 6 consecutive histidine residues (HHHHHH). The His tag is commonly expressed as a tag at either N-or C-terminal regions of recombinant proteins to allow isolation or purification by immobilized metal affinity chromatography.
What is his tag peptide used for?
Hexa His tag peptide is a 6 x His tag used as a metal binding site for the recombinant protein.
What does 6xHis stand for?
The 6xHis tag, also known as polyhistidine tag, His6 tag and/or hexa histidine tag, is an amino acid motif consisting of at least 6 histidine residues fused to the carboxyl (C-) or amino (N-) terminus of a target protein in transfected cells. This tag is most commonly used in the production…
What does His6 stand for?
A polyhistidine-tag is an amino acid motif in proteins that typically consists of at least six histidine ( His) residues, often at the N- or C-terminus of the protein. It is also known as hexa histidine-tag, 6xHis-tag, His6 tag, by the US trademarked name HIS TAG (US Trademark serial number 74242707),…