Human artificial chromosome containing human antibody λ light chain gene and non-human animal containing the human artificial chromosome capable of genetic transmission
The present invention relates to a human artificial chromosome which is genetically transmissible to the next generation with high efficiency and the method for using the same. More specifically, the present invention relates to: a human artificial chromosome in which an about 3.5 Mb to about 1 Mb region containing an antibody λ light chain gene derived from human chromosome 22 is bound to a chromosome fragment which is transmissible to a progeny through a germ line of a non-human animal, said chromosome fragment is derived from another human chromosome; a non-human animal carrying the human artificial chromosome and an offspring thereof; a method for producing the non-human animal; a method for producing a human antibody using the nonhuman animal or an offspring thereof; and a human antibody-producing mouse carrying the human artificial chromosome.
1. A method for producing a monoclonal antibody, which comprises:
immunizing with a desired antigen, a human antibody-producing mouse which carries (1) a human artificial chromosome in which a SC20 chromosome vector is linked to an about 1 Mb to about 3.5 Mb fragment of human chromosome 22 comprising an unrearranged human antibody λ light chain locus, and (2) an unrearranged human antibody κ light chain locus retained on a KCo5 transgene or on fragment W23 of human chromosome 2 (hCF(2-W23)), wherein at least both alleles of endogenous heavy chain locus and κ light chain locus are disrupted or inactivated, and which expresses a human antibody heavy chain comprising a human antibody Ig γ isotype, a human antibody κ light chain, and a human antibody λ light chain;
producing a hybridoma by fusing a mouse myeloma cell with the a spleen cell derived from the mouse; and
thereby producing a monoclonal antibody comprising human immunoglobulin heavy chains and light chains against the antigen.
2. The method of claim 1 , wherein the unrearranged human antibody κ light chain locus is inserted into a mouse chromosome.
3. The method of claim 1 , which further comprises:
isolating human heavy and light chain antibody genes from the hybridoma;
introducing the human heavy and light chain antibody genes into a cell capable of expressing the human heavy and light chain antibody genes;
culturing the cell under a condition capable of expressing the human heavy and light chain antibody genes; and
thereby producing a monoclonal antibody comprising heavy chains and light chains.
4. The method according to claim 3 , wherein the cell is an animal cell, a yeast cell or an insect cell.
5. The method of claim 1 , wherein the human antibody-producing mouse carries at least 40% of the variable region of the human antibody κ light chain.
6. The method of claim 1 , wherein the human antibody-producing mouse carries all the variable regions of the human antibody heavy chain, the human antibody κ light chain, and the human antibody λ light chain.
7. The method of claim 1 , wherein the human antibody κ light chain locus is retained on hCF(2-W23).
8. The method of claim 1 , wherein the human artificial chromosome is ΔΔHAC.
9. The method of claim 1 , wherein the human artificial chromosome is ΔHAC.
10. The method of claim 8 , wherein 100% of cells of the human antibody-producing mouse retain the human artificial chromosome.
11. The method of claim 1 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is from about 1.2 Mb to about 3 Mb.
12. The method of claim 1 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is from about 1.5 Mb to about 2.5 Mb.
13. The method of claim 1 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is 1.5 Mb.
14. The method of claim 1 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is 2.5 Mb.
15. A method for producing a monoclonal antibody, which comprises:
immunizing with a desired antigen, a human antibody-producing mouse which carries a human artificial chromosome in which a SC20 chromosome vector is linked to an about 1 Mb to about 3.5 Mb fragment of human chromosome 22 comprising an unrearranged human antibody λ light chain locus, wherein at least both alleles of endogenous heavy chain locus and κ light chain locus are disrupted or inactivated, and which expresses a human antibody heavy chain comprising a human antibody Ig γ isotype and a human antibody λ light chain;
producing a hybridoma by fusing a mouse myeloma cell with the a spleen cell derived from the mouse; and
thereby producing a monoclonal antibody comprising human immunoglobulin heavy chains and light chains against the antigen.
16. The method of claim 15 , which further comprises:
isolating human heavy and light chain antibody genes from the hybridoma;
introducing the human heavy and light chain antibody genes into a cell capable of expressing the human heavy and light chain antibody genes;
culturing the cell under a condition capable of expressing the human heavy and light chain antibody genes; and
thereby producing a monoclonal antibody comprising heavy chains and light chains.
17. The method according to claim 16 , wherein the cell is an animal cell, a yeast cell or an insect cell.
18. The method of claim 15 , wherein the human artificial chromosome is ΔΔHAC.
19. The method of claim 15 , wherein the human artificial chromosome is ΔHAC.
20. The method of claim 18 , wherein 100% of cells of the human antibody-producing mouse retain the human artificial chromosome.
21. The method of claim 15 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is from about 1.2 Mb to about 3 Mb.
22. The method of claim 15 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is from about 1.5 Mb to about 2.5 Mb.
23. The method of claim 15 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is 1.5 Mb.
24. The method of claim 15 , wherein the size of the fragment of human chromosome 22 comprising the unrearranged human antibody λ light chain locus is 2.5 Mb.