公开(公告)号：US10533204B2

公开(公告)日：2020-01-14

申请号：US15314862

申请日：2015-05-29

申请人： NATIONAL UNIVERSITY CORPORATION UNIVERSITY OF TOYAMA ,  SC WORLD, INC.

发明人： Hiroshi Hamana ,  Hiroyuki Kishi ,  Atsushi Muraguchi ,  Kiyomi Shitaoka

IPC分类号： C12P19/34 ,  C07K14/725 ,  C12Q1/6886 ,  A61K48/00

摘要： As a method for highly efficiently amplifying a TCR cDNA in a short period of time, there is provided a method for amplifying a T cell receptor (TCR) cDNA, which comprises the following step (1) and step (2): (1) the step of performing PCR by using at least one kind of the L primer mentioned below, the C primer 1 or UTR primer 1 mentioned below, and cDNA obtained from a single cell as the template to obtain an amplification product 1; an L primer of 30- to 60-nucleotide length comprising an adapter part of 15- to 25-nucleotide length, and a leader region-annealing part of 15- to 25-nucleotide length, which is ligated downstream from the adapter part, and can anneal to a part of a leader region containing a translation initiation codon, or an upstream part thereof, a C primer 1 of 15- to 25-nucleotide length, which can anneal to a part of a constant region, or a UTR primer 1 of 15- to 25-nucleotide length, which can anneal to a part of a 3′ untranslated region; (2) the step of performing PCR by using the adaptor primer mentioned below, the C primer 2 or UTR primer 2 mentioned below, and the amplification product 1 as the template to obtain an amplification product 2; an adapter primer of 15- to 25-nucleotide length, which can anneal to the adapter part of the amplification product 1, a C primer 2 of 15- to 25-nucleotide length, which can anneal to a part of the constant region existing upstream from the region to which the C primer 1 anneals, or a UTR primer 2 of 15- to 25-nucleotide length, which can anneal to a part of the 3′ untranslated region existing upstream from the region to which the UTR primer 1 anneals.

公开(公告)号：US20170183700A1

公开(公告)日：2017-06-29

申请号：US15314862

申请日：2015-05-29

申请人： SC WORLD, INC.

发明人： Hiroshi HAMANA ,  Hiroyuki KISHI ,  Atsushi MURAGUCHI ,  Kiyomi SHITAOKA

IPC分类号： C12P19/34 ,  C12Q1/68 ,  C07K14/725

摘要： As a method for highly efficiently amplifying a TCR cDNA in a short period of time, there is provided a method for amplifying a T cell receptor (TCR) cDNA, which comprises the following step (1) and step (2): (1) the step of performing PCR by using at least one kind of the L primer mentioned below, the C primer 1 or UTR primer 1 mentioned below, and cDNA obtained from a single cell as the template to obtain an amplification product 1; an L primer of 30- to 60-nucleotide length comprising an adapter part of 15- to 25-nucleotide length, and a leader region-annealing part of 15- to 25-nucleotide length, which is ligated downstream from the adapter part, and can anneal to a part of a leader region containing a translation initiation codon, or an upstream part thereof, a C primer 1 of 15- to 25-nucleotide length, which can anneal to a part of a constant region, or a UTR primer 1 of 15- to 25-nucleotide length, which can anneal to a part of a 3′ untranslated region; (2) the step of performing PCR by using the adaptor primer mentioned below, the C primer 2 or UTR primer 2 mentioned below, and the amplification product 1 as the template to obtain an amplification product 2; an adapter primer of 15- to 25-nucleotide length, which can anneal to the adapter part of the amplification product 1, a C primer 2 of 15- to 25-nucleotide length, which can anneal to a part of the constant region existing upstream from the region to which the C primer 1 anneals, or a UTR primer 2 of 15- to 25-nucleotide length, which can anneal to a part of the 3′ untranslated region existing upstream from the region to which the UTR primer 1 anneals.