IP Library › Granted Patent US 12,590,319
Granted Patent B2
US 12,590,319 · App. 17/847,021 · Granted Mar 31, 2026

Transposition of nucleic acid constructs into eukaryotic genomes with a transposase from amyelois

Inventors: Jeremy Minshull (Los Altos, CA); Sridhar Govindarajan (Los Altos, CA); Maggie Lee (San Jose, CA)
Assignee: DNA TWOPOINTO INC.
C12N15/85C12N2830/007
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Quick Facts
Patent No.
US 12,590,319
App. No.
17/847,021
Granted
Mar 31, 2026
Kind
B2
Abstract

The present invention provides polynucleotide vectors for high expression of heterologous genes. Some vectors further comprise novel transposons and transposases that further improve expression. Further disclosed are vectors that can be used in a gene transfer system for stably introducing nucleic acids into the DNA of a cell. The gene transfer systems can be used in methods, for example, gene expression, bioprocessing, gene therapy, insertional mutagenesis, or gene discovery.

Claims (15)

1 . A method of integrating a transposon into a eukaryotic cell, the method comprising (a) introducing into the cell a transposon comprising SEQ ID NO: 9 and SEQ ID NO: 10 as left and right ends respectively flanking a heterologous polynucleotide encoding a polypeptide; (b) introducing into the cell a transposase, the sequence of which is at least 90% identical to SEQ ID NO: 18, wherein the transposase transposes the transposon to produce a genome comprising SEQ ID NO: 9 and SEQ ID NO: 10 flanking the heterologous polynucleotide at left and right ends respectively; and (c) expressing the polypeptide from the heterologous polynucleotide within the genome.

2 . The method of claim 1 , wherein the polypeptide is a chimeric antigen receptor.

3 . The method of claim 1 , wherein the polypeptide comprises a chain of an antibody.

4 . The method of claim 1 , further comprising purifying the polypeptide.

5 . The method of claim 1 , further comprising incorporating the purified polypeptide into a pharmaceutical composition.

6 . The method of claim 1 , wherein the cell is a mammalian cell.

7 . The method of claim 1 , wherein the cell is a human cell.

8 . The method of claim 1 , wherein the cell is a rodent cell.

9 . The method of claim 1 , wherein the transposase is introduced as a polynucleotide encoding the transposase.

10 . The method of claim 9 , wherein the polynucleotide encoding the transposase is an mRNA.

11 . The method of claim 1 , wherein the transposase is introduced as a protein.

12 . The method of claim 1 , wherein the transposase has a sequence which comprises any of SEQ ID NOS: 19-35, or 51-170.

13 . The method of claim 1 , wherein the transposon further comprising a sequence at least 90% identical to SEQ ID NO: 13 immediately adjacent to the left transposon end and between the left transposon end and the heterologous polynucleotide and a sequence at least 90% identical to SEQ ID NO: 16 immediately adjacent to the right transposon end and between the right transposon end and the heterologous polynucleotide.

14 . The method of claim 1 , wherein the heterologous polynucleotide encodes two open reading frames, each operably linked to a separate promoter, one open reading frame encoding the polypeptide, and the other open reading frame a second polypeptide.

15 . The method of claim 14 , wherein the open reading frames encode heavy and light chains of an antibody.

Continuity (6)
Continuation 17339597 · Jun 4, 2021
Continuation 16842709 · Apr 7, 2020
Provisional Application 62990568 · Mar 17, 2020
Provisional Application 62873342 · Jul 12, 2019
Provisional Application 62831097 · Apr 8, 2019
Related Publication 20220411816A1 · Dec 29, 2022
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