Homologous recombination in multipotent adult progenitor cells
First Claim
Patent Images
1. A method of altering a first gene expression pattern in an isolated multipotent adult progenitor cell (MAPC) comprising:
- a) introducing into a MAPC an exogenous polynucleotide molecule, wherein the exogenous polynucleotide molecule comprises i) a targeting polynucleotide sequence homologous to a genomic DNA sequence of the MAPC and ii) a donor nucleotide sequence of interest; and
b) culturing the MAPC under conditions sufficient to homologously recombine the exogenous polynucleotide molecule, such that a resultant MAPC has a second expression pattern different than the first gene expression pattern.
5 Assignments
0 Petitions
Accused Products
Abstract
The invention relates to methods of altering gene expression by homologous recombination in a multipotent adult progenitor cell (MAPC). In particular, methods of producing a recombinant MAPC, of correcting a genetic defect in a mammal, of providing a functional and/or therapeutic protein to a mammal, and of transforming a MAPC are provided. MAPCs containing an erogenous DNA as well as recombinant MAPCs and their differentiated progeny are also provided.
29 Citations
20 Claims
-
1. A method of altering a first gene expression pattern in an isolated multipotent adult progenitor cell (MAPC) comprising:
-
a) introducing into a MAPC an exogenous polynucleotide molecule, wherein the exogenous polynucleotide molecule comprises i) a targeting polynucleotide sequence homologous to a genomic DNA sequence of the MAPC and ii) a donor nucleotide sequence of interest; and
b) culturing the MAPC under conditions sufficient to homologously recombine the exogenous polynucleotide molecule, such that a resultant MAPC has a second expression pattern different than the first gene expression pattern. - View Dependent Claims (2, 3, 4, 5)
-
-
6. A method of making recombinant multipotent adult progenitor cells (MAPCs) comprising:
-
a) culturing isolated MAPCs at low density;
b) nucleoporating the MAPC in the presence of an exogenous polynucleotide molecule, wherein the polynucleotide molecule comprises i) a targeting polynucleotide sequence homologous to a genomic DNA sequence of the MAPC and ii) optionally a DNA sequence encoding a gene product; and
c) culturing the MAPC obtained in b) under conditions sufficient to homologously recombine the exogenous DNA molecule, thereby making a recombinant MAPC. - View Dependent Claims (7, 8, 9)
-
-
10. A method of correcting a genetic defect in a mammal, wherein the defect is one or more defective nucleotide sequence(s) in the genome of the mammal that give(s) rise to defective gene expression, the method comprising:
-
a) culturing a MAPC from the mammal having the genetic defect;
b) introducing into the MAPC an exogenous DNA molecule, wherein the DNA molecule comprises i) a targeting DNA sequence homologous to a genomic DNA sequence of the MAPC and ii) one or more donor nucleotide sequence(s) necessary for correcting said genetic defect in said mammal, c) culturing the MAPC under conditions sufficient to homologously recombine the exogenous DNA molecule into the genome of the MAPC, thereby obtaining a genetically altered MAPC;
d) selecting said genetically altered MAPC; and
e) transplanting said genetically altered MAPC into the mammal;
wherein the selecting and transplanting can be done in any order or simultaneously. - View Dependent Claims (11, 12, 13, 14, 15, 16, 17)
-
- 18. A genetically altered MAPC comprising an exogenous polynucleotide molecule homologously recombined into the genome of a MAPC.
-
20. A method of expressing a functional gene product of interest in an isolated MAPC having a defective nucleotide sequence from which a functional gene product cannot be expressed, the method comprising:
-
a) introducing into the MAPC an exogenous DNA molecule, wherein the DNA molecule comprises i) a targeting DNA sequence homologous to a genomic DNA sequence of the MAPC and ii) a donor nucleotide sequence corresponding to the defective nucleotide sequence; and
b) culturing the MAPC under conditions sufficient to homologously recombine the exogenous DNA molecule into the genome of the MAPC, thereby expressing the functional gene product of interest in said MAPC.
-
Specification