Triple tag sequence and methods of use thereof
View Patent ↗The present disclosure relates to novel triple tag sequences that may comprise a 6× histidine tag, a c-myc tag and a V5 tag. The present disclosure also provides polynucleotides, proteins, vectors and host cells that comprise the triple tag sequence of the present disclosure, including libraries of such polynucleotides, proteins, vectors and host cells. The novel triple tag sequences of the present disclosure may be used in phage display vectors and phage libraries and in methods for detection, screening, capture, purification, quantitation, and/or recovery of proteins of interest to which they are linked. Proteins of interest include antibodies such as single chain antibodies, single chain antibodies, and Fab fragments of antibodies or peptides such as non-antibody peptides.
1. A method of screening a phagemid library of proteins of interest, wherein the library comprises a plurality of vectors which each comprise a polynucleotide coding for a protein of interest linked to a polynucleotide coding for a triple tag sequence, wherein the triple tag sequence comprises 6X histidine (SEQ ID NO: 1), c-myc (SEQ ID NO: 2) and V5 (SEQ ID NO: 3), said method comprising:
(a.) expressing a plurality of proteins of interest linked to the triple tag sequence from the plurality of vectors;
(b.) screening the proteins of interest linked to the triple tag sequence for binding to a binding partner of the protein of interest; and
(c.) recovering the proteins of interest linked to the triple tag sequence that are bound in step (b.) by capture with an antibody specific for V5, c-myc or 6X histidine,
wherein the triple tag sequence is HHHHHHGAAEQKLISEEDLKAAGKPIPNPLLGLDST (SEQ ID NO: 5), and wherein the protein of interest is a single chain antibody.