Binary hybrid mutational vectors
OwnerHoward B. Gamper, Eric B. Kmiec +1inventors
InventorsHoward B. Gamper, Eric B. Kmiec, Richard J. Bartlett
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Abstract
The invention concerns novel compounds used to make specific genetic alterations in the genome of target eukaryotic and prokaryotic cells, methods of their production and use, and kits containing the compounds. The compounds are two-strand duplex oligonucleobase compounds termed binary Hybrid Mutational Vectors (bHMV). A bHMV preferably contains a deoxyribo-type oligonucleobase mutator strand and a ribo-type oligonucleobase targeting strand. A bHMV is constructed to effect changes in a target gene via a combination of gene targeting and mismatch repair between the bHMV and the target gene.
Claims71
A binary hybrid mutational vector comprising an oligonucleobase targeting strand and an oligonucleobase mutator strand, wherein said targeting strand comprises:
- a) a 3′ end and a 5′ end;
- and
- b) a targeting strand homologous region comprising a sequence of nucleobases homologous to a target gene sequence;
- and wherein said mutator strand comprises:
- c) a 3′ end and a 5′ end;
- d) a complementary region comprising a sequence of nucleobases complementary to the targeting strand homologous region;
- and
- e) a mismatch region located within the complementary region.
A method of producing a binary hybrid mutational vector comprising the steps of:
- (a) providing a separately synthesized oligonucleobase targeting strand and oligonucleobase mutator strand, wherein said targeting strand comprises:
- 1) a 3′ end and a 5′ end;
- and
- 2) a targeting strand homologous region comprising a sequence of nucleobases homologous to a target gene sequence;
- and wherein said mutator strand comprises:
- 3) a3′ end and a 5′ end;
- 4) a complementary region comprising a sequence of nucleobases complementary to the targeting strand homologous region;
- and
- 5) a mismatch region located within the complementary region;
- and
- b) mixing said targeting strand and said mutator strand such that the mutator strand complementary region containing the mismatch region forms a duplex with the targeting strand homologous region.
An oligonucleobase set comprising an oligonucleobase targeting strand and a plurality of oligonucleobase mutator strands, wherein said targeting strand comprises:
- a) a 3′ end and a 5′ end;
- and
- b) a targeting strand homologous region comprising a sequence of nucleobases homologous to a target gene sequence;
- and wherein each mutator strand comprises:
- c) a 3′ end and a 5′ end;
- dd) a complementary region comprising a sequence of nucleobases complementary to the targeting strand homologous region;
- and
- e) a unique mismatch region located within the complementary region.
Dependencies are inferred from the claim text.