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Fluorescence-based, Nonradioactive Method for Efficient Detection of the Pentanucleotide Repeat (TTTTA)_n Polymorphism in the Apolipoprotein(a) Gene

¹ Department of Medicine, Columbia University, New York, NY 10032.

² Department of Community and Family Medicine, University of Rochester, Rochester, NY 14642.

³ Bassett Research Institute, Cooperstown, NY 13326.

^aAddress correspondence to this author at: Department of Medicine, Room PH 10-305, Columbia University, 630 West 168th St., New York, NY 10032. Fax 212-305-3213; e-mail lfb9{at}columbia.edu.

Background: The apolipoprotein(a) [apo(a)] gene is a major predictor of plasma lipoprotein(a) concentrations, an independent risk factor for cardiovascular disease. The apo(a) gene contains a pentanucleotide repeat (PNR) polymorphism, 1.4 kb upstream from the apo(a) gene reading frame. This polymorphism has been suggested to be important in control of apo(a) gene expression.

Methods: We developed a fluorescence-based, nonradioactive procedure to detect the PNR polymorphism. After amplification of the polymorphism by PCR, the respective PCR products were separated by denaturing polyacrylamide gel electrophoresis and detected using a 3'-end fluorescently labeled oligonucleotide as a probe. We used the method to characterize the PNR polymorphism pattern in 313 individuals, 195 Caucasians and 118 African Americans. The new method efficiently separated DNAs corresponding to the different PNR repeats.

Results: Among both ethnic groups, alleles containing eight PNRs were most common. Smaller PNRs were more common among African Americans, and larger PNRs were more common among Caucasians.

Conclusions: We developed a nonradioactive technique that separates the PNR polymorphism in the apo(a) gene and can be used in other studies involving closely sized polymorphisms.

The following articles in journals at HighWire Press have cited this article:

				J. Rubin, H. J. Kim, T. A. Pearson, S. Holleran, R. Ramakrishnan, and L. Berglund Apo[a] size and PNR explain African American-Caucasian differences in allele-specific apo[a] levels for small but not large apo[a] J. Lipid Res., May 1, 2006; 47(5): 982 - 989. [Abstract] [Full Text] [PDF]

				L. Berglund and R. Ramakrishnan Lipoprotein(a): An Elusive Cardiovascular Risk Factor Arterioscler. Thromb. Vasc. Biol., December 1, 2004; 24(12): 2219 - 2226. [Abstract] [Full Text] [PDF]