Purification of STR-multiplex-amplified microsamples can enhance signal intensity in capillary electrophoresis.
In this study, the effect of sample purification on total signal intensities of samples amplified with genRES MPX-2 (nine-locus multiplex) prior to capillary electrophoretic analysis has been investigated. Sample purification with the Qiaquick PCR purification kit led to an increase of the relative fluorescent signal intensity by a factor of 3.8 +/- 0.8. In contrast, the application of larger sample volumes led to a decrease of signal intensities from 20% to 80%, depending on whether the samples were purified or not. In addition, increase of injection time showed a linear increase of signal intensity between 3 s and 10 s. Increasing the number of PCR cycles from 30 to 33 also led to a significant increase of signal intensities. Nevertheless, this increase greatly depended on the fragment lengths and was sometimes accompanied by the appearance of non-specific signals. In combination, optimisation of sample preparation and increase of injection time may intensify signals up to 12-fold, thereby increasing the overall sensitivity of the assay. This may be of special interest for forensic analysis of microspecimens containing limited amounts of DNA.
Lederer
T
T
Institut für Rechtsmedizin, Universität Erlangen-Nürnberg, Universitätsstrasse 22, 91054 Erlangen, Germany. thomas.lederer@recht.imed.uni-erlangen.de
Braunschweiger
G
G
Betz
P
P
Seidl
S
S
D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D019075:Electrophoresis, Capillary / Q000379:methods*; D005554:Forensic Medicine / Q000379:methods*; D006801:Humans; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D020080:Tandem Repeat Sequences / Q000235:genetics*
D004247:DNA