Methods for preparing cell lines that contain plant artificial chromosomes, methods for preparation of plant artificial chromosomes, methods for targete d insertion of heterologous DNA into plant artificial chromosomes, and methods for delivery of plant chromosomes to selected cells and tissues are provided . In particular, plant artificial chromosomes that are substantially composed of repeated nucleic acid units of varying amounts of heterochromatin and euchromatin are provided. Also provided are methods of using plant and anima l artificial chromosomes in the production of valuable transgenic plants. Methods for identifying plant genes encoding particular traits using artificial chromosomes and for producing an acrocentric plant chromosome are also provided.