US 11,702,765 B2
Recombinant fusion proteins and libraries from immune cell repertoires
David Scott Johnson, San Francisco, CA (US); Adam Adler, Belmont, CA (US); and Rena Mizrahi, Pacifica, CA (US)
Assigned to GigaGen, Inc., San Carlos, CA (US)
Filed by GigaGen, Inc., San Carlos, CA (US)
Filed on May 28, 2020, as Appl. No. 16/886,612.
Application 15/156,214 is a division of application No. 14/734,953, filed on Jun. 9, 2015, granted, now 9,422,547, issued on Aug. 23, 2016.
Application 16/886,612 is a continuation of application No. 16/399,759, filed on Apr. 30, 2019, granted, now 10,689,641.
Application 16/399,759 is a continuation of application No. 15/156,214, filed on May 16, 2016, abandoned.
Prior Publication US 2020/0291390 A1, Sep. 17, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. C40B 40/08 (2006.01); C07K 16/00 (2006.01); C12N 15/10 (2006.01); C07K 14/725 (2006.01); C12Q 1/6876 (2018.01); C40B 50/06 (2006.01)
CPC C40B 40/08 (2013.01) [C07K 14/7051 (2013.01); C07K 16/00 (2013.01); C12N 15/1037 (2013.01); C12N 15/1075 (2013.01); C12N 15/1093 (2013.01); C12Q 1/6876 (2013.01); C07K 2317/14 (2013.01); C07K 2317/622 (2013.01); C40B 50/06 (2013.01)] 10 Claims
OG exemplary drawing
 
1. A kit for producing a library of constructs for expression of thousands of full-length recombinant immunoglobulins, the kit comprising:
a. a plurality of at least 10,000 unique recombinant fusion polynucleotides, each comprising a first polynucleotide and a second polynucleotide and a linker polynucleotide,
each first polynucleotide encoding a heavy chain variable domain from a cognate pair from a single isolated mammalian B cell and an IgG heavy chain constant domain fragment; and
each second polynucleotide encoding a light chain variable domain from the cognate pair from the single isolated mammalian B cell and an IgK light chain constant domain; and
the linker polynucleotide linking the first and second polynucleotides and comprising a restriction site;
b. a third recombinant polynucleotide, not operationally linked to the recombinant fusion polynucleotides, comprising a first promoter sequence and a sequence encoding a portion of the IgG heavy chain constant domain, wherein the third recombinant polynucleotide comprises a first end sequence and a second end sequence, each overlapping with a first or second end sequence of each recombinant fusion polynucleotide;
c. a fourth recombinant polynucleotide, not operationally linked to the recombinant fusion polynucleotides or the third recombinant polynucleotide, comprising a second promoter sequence or a translational skip site; and
d. a mixture for Gibson assembly.