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Current Topics in Analytical Chemistry   Volumes    Volume 6 
Abstract
The use of microchip electrophoresis in Short Tandem Repeat (STR) DNA analysis and its applicability for developing methods for Low Copy Number (LCN) DNA profiling
Henrietta Margolis-Nunno, Ewelina Bajda, Miriam Cohen, Lawrence Kobilinsky
Pages: 25 - 39
Number of pages: 15
Current Topics in Analytical Chemistry
Volume 6 

Copyright © 2006 Research Trends. All rights reserved

ABSTRACT
 
The Agilent 2100 Bioanalyzer, which separates, sizes and quantifies DNA using microfluidic technology, was evaluated for its usefulness in short tandem repeat (STR) low copy number (LCN) DNA analysis. Studies employing unlabeled commercial STR triplex primers and genomic DNA with known STR alleles, demonstrate that the Bioanalyzer is capable of distinguishing heterozygous STR alleles that are 7-8 base pairs apart as two distinct peaks, and homozygous alleles as a single sharp peak. Studies using STR fragments of known sizes and DNA standards and ladders, show that although certain STR loci consistently show relatively larger sizing errors than others, all results are reproducible with low values for the coefficient of variation and all are within an error range of 5%. In studies using DNA concentrations that included those found in LCN DNA, Bioanalyzer profiles of PCR products show DNA concentration dependent differences and a ten-fold increase in sensitivity when the number of PCR cycles is increased from 30 to 34. In addition, the type of stochastic variation that is common to the amplification of LCN DNA is easily visible with the Bioanalyzer; allele or locus drop out and imbalance and allele drop in were all seen when LCN DNA concentrations of 25 and  2.5 pg were amplified for 34 cycles. Thus, although the accuracy of the Bioanalyzer is not sufficient for STR allele identification, its allele resolution, ease of use, speed, economy and small sample requirement, make it very useful for screening amplified product for preliminary research and technique development in the analysis of STRs in LCN DNA samples.
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