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    Methodology for detecting trace amounts of microchimeric DNA from peripheral murine white blood cells by real-time PCR

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    Genre
    Journal Article
    Date
    2003-12-01
    Author
    Artlett, CM
    Dito, CG
    Christner, PJ
    Permanent link to this record
    http://hdl.handle.net/20.500.12613/5662
    
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    DOI
    10.1251/bpo51
    Abstract
    Real-time PCR methodology can successfully quantitate microchimeric cell populations at a concentration of 100 microchimeric cells/100,000 host cells; however, it has not been successful in quantitating DNA from trace numbers of microchimeric white blood cells which we reported are present in murine peripheral blood at a concentration as low as 2/100,000 host cells. We report methodology using primers for a portion of the H2-kb murine histocompatibility sequence, specific for the C57BL/6J mouse. When these primers were used in the presence of 11,000 μM primer, a 20-fold increase in the median manufacturer's recommended concentration, the assay could be optimized to detect 34 pg of C57BL/6J DNA in a background of 2.5 μg of carrier BALB/cJ DNA (1/100,000). These conditions resulted in a detection limit half as sensitive as that found when no carrier DNA was present.
    Citation to related work
    Springer Science and Business Media LLC
    Has part
    Biological Procedures Online
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    ae974a485f413a2113503eed53cd6c53
    http://dx.doi.org/10.34944/dspace/5644
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