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

被引：3

作者：

Carol M. Artlett

C. Gennaro Dito

Paul J. Christner

机构：

[1] Division of Rheumatology, Thomas Jefferson University, Philadelphia, PA 19107

来源：

Biological Procedures Online | 2003年 / 5卷 / 1期

关键词：

DNA; Leukocytes; Mice; Polymerase chain reaction; T-lymphocytes;

D O I：

10.1251/bpo51

中图分类号：

学科分类号：

摘要：

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.

引用

页码：103 / 107

页数：4

共 10 条

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