Mammalian artificial chromosomes
AssigneeBiologic Research Center of the Hungarian Academy of Sciences
InventorGyula Hadlaczky
Life of this patent
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Abstract
Non-human cell lines are disclosed which contain functional centromeres comprising human DNA sequences linked to a dominant marker gene. The centromeres are carried on stable chromosomes which carry no centromeres other than those comprising human DNA sequences. The cell lines can be used to isolate the chromosomes as well as for use in inserting genes into mammalian cells. Methods are taught for generating such cell lines from cell lines carrying dicentric chromosomes.
Claims12
A method of producing a cell line which comprises a functioning centromere comprising human DNA sequences, wherein the centromere is present on a chromosome, wherein all of the centromeres of said chromosome comprise human sequences, comprising:
- fusing cells of a cell line deposited at the European Collection of Animal Cell Culture (ECACC) under accession no. 90051001 with CHO K-20 cells to form fused cell hybrids;
- screening said fused cell hybrids or progeny for cells which are devoid of a dicentric chromosome but which contain a centromere which comprises human DNA.
A method of producing a cell line which comprises a functioning centromere comprising human DNA sequences, wherein the centromere is present on a chromosome, wherein all of the centromeres of said chromosome comprise human sequences, comprising:
- growing cells of a cell line deposited at the European Collection of Animal Cell Culture (ECACC) under accession no. 90051001 in a culture medium comprising a selective agent for an aminoglycoside-3' phosphotranferase-II gene, said selective agent present in the culture medium in an amount greater than ten times the amount which kills 50% of the cells;
- selecting cell which survive in said culture medium;
- screening the surviving cells or their progeny for cells which are devoid of a dicentric chromosome.
A non-human cell line which comprises a functioning centromere derived from a cell line which is deposited at the European Collection of Animal Cell Cultures (ECACC) under accession no. 90051001, wherein the centromere is present on a chromosome wherein all of the centromeres of said chromosome comprise cloned human sequences.
Dependencies are inferred from the claim text.