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Last Updated: March 29, 2024

Claims for Patent: 9,085,767


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Summary for Patent: 9,085,767
Title:Enhancer-containing gene trap vectors for random and targeted gene trapping
Abstract: The present invention relates to a novel class of gene trap vector (enhanced gene trap vectors, eGTV) for efficiently identifying silent or weakly expressed target genes in mammalian genomes, methods of their production and methods for identifying and mutating target genes by using the enhanced gene trap vectors. The gene trap vectors of the present invention can also be used for inducing the expression of silent genes and enhancing the expression of weakly expressed genes. The use of the enhanced gene trap vectors for creating transgenic organisms to identify gene function and to validate pharmaceutical compounds prior to clinical applications is a further aspect of the present invention.
Inventor(s): Von Melchner; Harald (Kronberg/Taunus, DE), Schnutgen; Frank (Alzenau, DE), Wurst; Wolfgang (Munchen, DE), Ruiz; Patricia (Berlin, DE)
Assignee: FRANKGEN BIOTECHNOLOGIE AG (Kronberg, DE) GSF FORSCHUNGSZENTRUM FUR UMWELT UND GESUNDHEIT GMBH (Neuherberg, DE) MPG MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN E.V. (Munich, DE)
Application Number:11/720,227
Patent Claims:1. A gene trap vector comprising a gene disruption and selection cassette (GDSC) and one or more non-viral, cell-type-specific enhancer elements that are active in mammalian cells, wherein said GDSC comprises a promoterless reporter gene and/or a promoterless selectable marker gene flanked upstream by a 3' splice acceptor (SA) site and downstream by a transcriptional termination polyA sequence.

2. The gene trap vector of claim 1, wherein the vector is a plasmid.

3. The gene trap vector of claim 1, wherein said one or more non-viral, cell-type-specific enhancer elements (i) are located upstream of the GDSC, and/or (ii) are located downstream of the GDSC.

4. The gene trap vector of claim 1, wherein the one or more non-viral, cell-type-specific enhancer elements contain at least one binding site for a transcription activating factor.

5. The gene trap vector of claim 1, wherein the one or more non-viral, cell-type-specific enhancer elements contain binding sites that bind transcription activation factors in a sequence-specific manner.

6. The gene trap vector of claim 5, wherein the binding sites are arranged as tandem repeats.

7. The gene trap vector of claim 1, wherein the one or more non-viral, cell-type-specific enhancer elements are selected from the group consisting of hormone responsive elements and transcription factor binding elements.

8. The gene trap vector of claim 1, wherein the one or more non-viral, cell-type-specific enhancer elements are tandem repeats that comprise a transcription factor binding site selected from the group consisting of NF-kB, Oct2, Oct3, and Oct4 transcription factor binding sites.

9. The gene trap vector of claim 8, wherein the transcription factor binding sites are the Oct-4 transcription factor binding sites.

10. The gene trap vector of claim 9, wherein the Oct-4 transcription factor binding site are inserted between two homotypic or heterotypic site-specific recombination targets.

11. The gene trap vector of claim 1, wherein the reporter gene is a R-galactosidase neomycinphosphotransferase fusion gene.

12. The gene trap vector of claim 1, further comprising a puromycin resistance gene inserted upstream or downstream of the GDSC.

13. The gene trap vector of claim 12, wherein the puromycin resistance gene is in direct or inverse transcriptional orientation relative to the GDSC.

14. The gene trap vector of claim 1, further comprising site-specific recombinase recognition elements that are oriented in opposing directions and flank the GDSC and the one or more enhancer elements such that inversion of the GDSC and the one or more enhancer element is mediated in the presence of the site-specific recombinase.

15. The gene trap vector of claim 14, wherein the recombinase is a Cre or a Flp recombinase.

16. The gene trap vector of claim 1, wherein the vector comprises a 5' and a 3' nucleotide sequence homologous to the 5' and the 3' sequence of an intron of a target gene, wherein the nucleotide sequences flank the GDSC and the one or more non-viral, cell-type-specific enhancer elements and further mediate homologous recombination at the intron.

17. A mammalian cell comprising the gene trap vector of claim 1.

18. A retroviral gene trap vector comprising a GDSC and one or more cell-type-specific enhancer elements that are active in mammalian cells, wherein said GDSC comprises a promoterless reporter gene and/or a promoterless selectable marker gene flanked upstream by a 3' splice acceptor (SA) site and downstream by a transcriptional termination polyA sequence.

19. The retroviral gene trap vector of claim 18, which is eFIip3ROSApgeo or eFIip6ROSApgeo having the nucleotide sequence of SEQ ID Nos:4 and 5, respectively.

Details for Patent 9,085,767

Applicant Tradename Biologic Ingredient Dosage Form BLA Approval Date Patent No. Expiredate
Merck Sharp & Dohme Corp. INTRON A interferon alfa-2b For Injection 103132 06/04/1986 ⤷  Try a Trial 2024-11-26
Merck Sharp & Dohme Corp. INTRON A interferon alfa-2b For Injection 103132 ⤷  Try a Trial 2024-11-26
Merck Sharp & Dohme Corp. INTRON A interferon alfa-2b Injection 103132 ⤷  Try a Trial 2024-11-26
>Applicant >Tradename >Biologic Ingredient >Dosage Form >BLA >Approval Date >Patent No. >Expiredate

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