Method for targeting a polypeptide onto cellular surface

A construct for introducing into a host cell for localizing a heterogeneous polypeptide on a cell surface, comprising one nucleotide sequence selected from the group consisting of the following (A) to (B):

• (A) a nucleotide sequence encoding an amino acid sequence selected from the group consisting of the following (a) to (b):
• (a) the amino acid sequence shown in SEQ ID NO:
• 2;
• and
• (b) an amino acid sequence with a mutation in SEQ ID NO:
• 2 selected from the group consisting of the following (1) to (34):
• (1) substitution of an amino acid residue at position 1 to T,
• (2) substitution of an amino acid residue at position 2 to t,
• (3) substitution of an amino acid residue at position 3 to P or T,
• (4) substitution of an amino acid residue at position 4 to T,
• (5) substitution of an amino acid residue at position 5 to Q,
• (6) substitution of an amino acid residue at position 6 to A,
• (7) substitution of an amino acid residue at position 7 to F or P,
• (8) substitution of an amino acid residue at position 8 to T,
• (9) substitution of an amino acid residue at position 9 to P,
• (10) substitution of an ammo acid residue at position 11 to L, S or P,
• (11) substitution of an ammo acid residue at position 12 to N, S or A,
• (12) substitution of an amino acid residue at position 13 to S,
• (13) substitution of an amino acid residue at position 15 to P,
• (14) substitution of an amino acid residue at position 17 to S or P,
• (15) substitution of an amino acid residue at position 18 to P, T or A,
• (16) substitution of an amino acid residue at position 19 to P,
• (17) substitution of an amino acid residue at position 20 to T,
• (18) substitution of an amino acid residue at position 21 to S or I,
• (19) substitution of an amino acid residue at position 23 to S or T,
• (20) substitution of an amino acid residue at position 24 to P or A,
• (21) substitution of an amino acid residue at position 25 to T,
• (22) substitution of an amino acid residue at position 26 to I,
• (23) substitution of an amino acid residue at position 27 to S,
• (24) substitution of an amino acid residue at position 28 to S,
• (25) substitution of an amino acid residue at position 30 to T or Q,
• (26) substitution of an amino acid residue at position 31 to N, S, T or A,
• (27) substitution of an amino acid residue at position 32 to F or C,
• (28) substitution of an amino acid residue at position 33 to R,
• (29) substitution of an amino acid residue at position 34 to T,
• (30) substitution of an amino acid residue at position 35 to E or T,
• (31) substitution of an amino acid residue at position 36 to T,
• (32) deletion of a sequence consisting of amino acids at positions 1 to 8,
• (33) deletion of a sequence consisting of amino acids at positions 1 to 7, and
• (34) deletion of a sequence consisting of amino acids at positions 32 to 36;
• and
• (B) a nucleotide sequence of a nucleic acid that hybridizes to an antisense sequence of the nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:
• 1 under conditions of carrying out incubation at 65° C., overnight in a solution containing 6×SSC, 0.5% SDS, 5×Denhardt's, and 100 μg/ml herring sperm DNA, and carrying out washing using a buffer with 2×SSC at a temperature of 25° C. below a Tm value of the nucleic acid used,
• wherein a polypeptide encoded by the nucleotide sequence has an activity for targeting a heterogeneous polypeptide to a surface of cytoplasmic membrane, and
• wherein a nucleic acid encoding a heterogeneous polypeptide to be expressed is operably linked downstream to a .nucleotide sequence selected from the group consisting of said (A) to (B) via a nucleotide sequence for operably linking the nucleotide encoding a heterogeneous polypeptide.

A pair of primers consisting of an oligonulceotide that hybridizes to a nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:

• 1 under stringent conditions, wherein the oligonucleotide consists of 17 to 50 nucleotides in length;
• and an oligonucleotide that hybridizes to an antisense nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:1 under stringent conditions, wherein the oligonucleotide consists of 17 to 50 nucleotides in length;
• and
• wherein said stringent conditions are the conditions for carrying out incubation at a temperature of 25° C. below a Tm value of said pair of primers overnight in a solution containing 6×SSC, 0.5% SDS, 5×Denhardt's, and 100 μg/ml herring sperm DNA, and carrying out washing using a buffer with 2×SSC at a temperature of 25° C. below a Tm value of said pair of primers.

A kit used for detecting a nucleic acid encoding a polypeptide having an activity for targeting a heterogeneous polypeptide to a surface of cytoplasmic membrane, comprising an oligonucleotide probe that hybridizes to a nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:

• 1 under stringent conditions, wherein the oligonucleotide probe consists of 18 to 150 nucleotides in length and/or a pair of primers consisting of an oligonucleotide that hybridizes to a nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:1 under stringent conditions, wherein the oligonucleotide consists of 17 to 50 nucleotides in length;
• and an oligonucleotide that hybridizes to an antisense nucleic acid consisting of the nucleotide sequence shown in SEQ ID NO:
• 1 under stringent conditions, wherein the oligonucleotide consists of 17 to 50 nucleotides in length;
• and wherein said stringent conditions are the conditions for carrying out incubation at a temperature of 25° C. below a Tm value of said oligonucleotide probe or said pair of primers overnight in a solution containing 6×SSC, 0.5% SDS, 5×Denhardt's, and 100 μg/ml herring sperm DNA, and carrying out washing using a buffer with 2×SSC at a temperature of 25° C. below a Tm value of said oligonucleotide probe or said pair of primers.