Artificial recombinant chromosome and use thereof

A method for producing a cell including one or more artificial recombinant chromosomes, the method comprising

• i) preparing a first targeted cell and a second targeted cell,
• wherein the first targeted cell comprises a first targeted chromosome, and the second targeted cell comprises a second targeted chromosome,
• wherein the first targeted chromosome includes a first part, a first recombinase recognition sequence (a first RRS), a first fragment, a second recombinase recognition sequence (a second RRS) and a second part,
• herein, the first part includes one of telomere ends located on both sides of the first targeted chromosome, and the second part includes the other one of telomere ends located on both sides of the first targeted chromosome,
• herein, the first fragment is located between the first RRS and the second RRS,
• wherein the second targeted chromosome includes a third part, a third recombinase recognition sequence (a third RRS), a second fragment, a fourth recombinase recognition sequence (a fourth RRS) and a fourth part,
• herein, the third part includes one of telomere ends located on both sides of the second targeted chromosome, and the fourth part includes the other one of telomere ends located on both sides of the second targeted chromosome,
• herein, the second fragment is located between the third RRS and the fourth RRS;
• ii) producing one or more microcells using the second targeted cell,
• wherein the one or more microcells comprise the second targeted chromosome or a fragment thereof,
• herein, the fragment of the second targeted chromosome includes the third RRS, the second fragment and the fourth RRS;
• iii) producing a fusion cell using the first targeted cell and the one or more microcells,
• wherein the fusion cell comprises the first targeted chromosome and the second targeted chromosome, or the first targeted chromosome and the fragment of the second targeted chromosome;
• and
• iv) producing a cell including an artificial recombinant chromosome by treating the fusion cell with site specific recombinase (SSR),
• wherein the SSR induces a recombination by recognizing a pairing of the first RRS present in the first targeted chromosome and the third RRS present in the second targeted chromosome, and a pairing of the second RRS present in the first targeted chromosome and the fourth RRS present in the second targeted chromosome,
• herein, the first fragment present in the first targeted chromosome is exchanged for the second fragment present in the second targeted chromosome by the recombination,
• thereby, a first artificial recombinant chromosome including the first part, the second fragment and the second part is produced.

A method for making a transgenic non-human animal using a cell including one or more artificial recombinant chromosome, the method comprising

• i) preparing a first targeted cell and a second target cell,
• wherein the first targeted cell is an embryonic stem cell,
• wherein the first targeted cell comprises a first targeted chromosome, and the second targeted cell comprises a second targeted chromosome,
• wherein the first targeted chromosome includes a first part, a first recombinase recognition sequence (a first RRS), a first fragment, a second recombinase recognition sequence (a second RRS) and a second part,
• herein, the first part includes one of telomere ends located on both sides of the first targeted chromosome, and the second part includes the other one of telomere ends located on both sides of the first targeted chromosome,
• herein, the first fragment is located between the first RRS and the second RRS,
• wherein the second targeted chromosome includes a third part, a third recombinase recognition sequence (a third RRS), a second fragment, a fourth recombinase recognition sequence (a fourth RRS) and a fourth part,
• herein, the third part includes one of telomere ends located on both sides of the second targeted chromosome, and the fourth part includes the other one of the telomere ends located on both sides of the second targeted chromosome,
• herein, the second fragment is located between the third RRS and the fourth RRS;
• ii) producing one or more microcells using the second targeted cell,
• wherein the one or more microcells comprises the second targeted chromosome or a fragment thereof,
• herein, the fragment of the second targeted chromosome includes the third RRS, the second fragment and the fourth RRS;
• iii) producing a fusion cell using the first targeted cell and the one or more microcells,
• wherein the fusion cell comprises the first targeted chromosome and the second targeted chromosome, or the first targeted chromosome and the fragment of the second targeted chromosome;
• iv) producing a cell including an artificial recombinant chromosome by treating the fusion cell with a site specific recombinase (SSR),
• wherein the SSR induces a recombination by recognizing a pairing of the first RRS present in the first targeted chromosome and the third RRS present in the second targeted chromosome, and a pairing of the second RRS present in the first targeted chromosome and the fourth RRS present in the second targeted chromosome,
• herein, the first fragment present in the first targeted chromosome is exchanged for the second fragment present in the second targeted chromosome,
• thereby, a first artificial recombinant chromosome including the first part, the second fragment and the second part is produced;
• and
• v) implanting a chimeric blastocyst comprising the first artificial recombinant chromosome in a surrogate mother's uterus to produce an offspring,
• herein, the chimeric blastocyst is produced by injecting the cell including the first artificial recombinant chromosome into a blastocyst.