クロレラ染色体基本構造ユニットの解析とその再構成による人工染色体のモデル化
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The future growth of plant biotechnology will be built upon a better understanding of complex, multigene traits, including the pathways of natural-product biosynthesis and the mechanisms of resistance against environmental stresses, along with a better integration of biochemistry, physiology, genetics, and developmental and cellular biology. Development of new tools and methods for the transfer of multiple genes (sometimes in the order of a hundred) to plants will be required for various purposes. The plant artificial chromosome (PAC) promises to be a powerful candidate as a next-generation vector in plant transformation. Learning from the success of YACs (yeast artificial chromosomes) in the cloning of huge human DNAs, it is clear that a minimum of three types of chromosome elements are needed to construct a linear, mitotically stable artificial chromosome : a centromere, telomeres, and origins of replication. The natures of the corresponding elements in plant chromosomes are, however, less well understood. To obtain an experimental model for molecular and structural analyses, we screened a variety of organisms by pulsed-field gel electrophoresis for the smallest plant-type chromosome. Species of the unicellular green alga Chlorella were found to contain very small genomes of approximately 39 Mbp, almost a quarter the size of the Arabidopsis genome. In the case of Chlorella vulgaris C-169,16 chromosomes ranging from 980 Kbp to 4.0 Mbp in size could be resolved by CHEF gel electrophoresis under ordinary conditions. The smallest chromosome of this strain (chromosome I, 980 Kbp) would serve as an excellent experimental subject for structural analysis because it can be routinely isolated easily and in quantity. With this chromosomal DNA, a set of overlapping cosmid clones (a contig) has been established. Telomeres obtained from this chromosome consist of 5'-TTTAGGG repeats running from the replication origins and centromeric components isolated as curved DNA from C-169 chromosome I were characterized by sequencing and localizing on the chromosome. By combining the Chlorella chromosomal structural elements thus obtained, various forms of artificial chromosomes can be now designed to be introduced and stably maintained in plants.
- 1998-05-25
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