Infrared fluorescent automated detection of thirteen short tandem repeat polymorphisms and one gender-determining system of the CODIS core system.
We used an infrared (IR) automated fluorescence monolaser sequencer for the analysis of 13 autosomal short tandem repeat (STR) systems (TPOX, D3S1358, FGA, CSF1PO, D5S818, D7S820, D8S1179, TH01, vWA, D13S317, D16S359, D18S51, D21S11) and the X-Y homologous gene amelogenin system. These two systems represent the core of the combined DNA index systems (CODIS). Four independent multiplex reactions, based on the polymerase chain reaction (PCR) technique and on the direct labeling of the forward primer of every primer pair, with a new molecule (IRDye800), were set up, permitting the exact characterization of the alleles by comparison with ladders of specific sequenced alleles. This is the first report of the whole analysis of the STRs of the CODIS core using an IR automated DNA sequencer. The protocol was used to solve paternity/maternity tests and for population studies. The electrophoretic system also proved useful for the correct typing of those loci differing in size by only 2 bp. A sensibility study demonstrated that the test can detect an average of 10 pg of undegraded human DNA. We also performed a preliminary study analyzing some forensic samples and mixed stains, which suggested the usefulness of using this analytical system for human identification as well as for forensic purposes.
Ricci
U
U
Department of Pediatrics, University of Florence, Italy. uzielli_ml@unifi.it
Sani
I
I
Guarducci
S
S
Biondi
C
C
Pelagatti
S
S
Lazzerini
V
V
Brusaferri
A
A
Lapini
M
M
Andreucci
E
E
Giunti
L
L
Giovannucci Uzielli
M L
ML
D001331:Automation; D004247:DNA / Q000032:analysis; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D007211:Indoles / Q000737:chemistry; D007259:Infrared Rays; D018598:Minisatellite Repeats; D015394:Molecular Structure; D021141:Nucleic Acid Amplification Techniques; D010334:Paternity; D011110:Polymorphism, Genetic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000379:methods*; D013698:Templates, Genetic
D005456:Fluorescent Dyes; C427728:IRDye800; D007211:Indoles; D004247:DNA