Construction and characterization of a yeast artificial chromosome library containing seven haploid human genome equivalents

Hans M. Albertsen, Hadi Abderrahim, Howard M. Cann, Jean Dausset, Denis L.E. Paslier, Daniel Cohen

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    490 Citations (Scopus)

    Abstract

    Prior to constructing a library of yeast artificial chromosomes (YACs) containing very large human DNA fragments, we performed a series of preliminary experiments aimed at developing a suitable protocol. We found an inverse relationship between YAC insert size and transformation efficiency. Evidence of occasional rearrangement within YAC inserts was found resulting in clonally stable internal deletions or clonally unstable size variations. A protocol was developed for preparative electrophoretic enrichment of high molecular mass human DNA fragments from partial restriction digests and ligation with the YAC vector in agarose. A YAC library has been constructed from large fragments of DNA from an Epstein-Barr virus-transformed human lymphoblastoid cell line. The library presently contains 50,000 clones, 95% of which are greater than 250 kilobase pairs in size. The mean YAC size of the library, calculated from 132 randomly isolated clones, is 430 kilobase pairs. The library thus contains the equivalent of approximately seven haploid human genomes.

    Original languageEnglish
    Pages (from-to)4256-4260
    Number of pages5
    JournalProceedings of the National Academy of Sciences of the United States of America
    Volume87
    Issue number11
    DOIs
    Publication statusPublished - 1 Jan 1990

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    Keywords

    • Physical mapping
    • Preparative pulsed-field electrophoresis

    ASJC Scopus subject areas

    • General

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