The analysis of short tandem repeats - An Introduction to Forensic Genetics - page 70

Detection of STR polymorphisms

After STR polymorphisms have been amplified using PCR, the length of the products
must be measured precisely - some STR alleles differ by only 1 bp. Gel electrophore-
sis of the PCR products through denaturing polyacrylamide gels can be used to
separate DNA molecules between 20 and 500 nucleotides long with single base pair
resolution [30]. Early systems detected the PCR products after electrophoresis on
polyacrylamide slab-gels using silver staining [31, 32], but this limited the number
of loci that could be incorporated into the multiplexes because the allelic size ranges
of the different loci could not overlap. To overcome this limitation, fluorescence
labelling of PCR products followed by multicolour detection has been adopted by
the forensic community. A series of fluorescent dyes has been developed that can
be covalently attached to the 5 end of one of the PCR primers in each primer pair
and detected real-time during electrophoresis. Up to five different dyes can be used
in a single analysis, which allows for considerable overlap of loci (Figure 6.3). The
electrophoresis platforms have evolved from systems based on slab-gels to capillary
electrophoresis (CE), which use a narrow glass tube filled with an entangled polymer

AmpFlSTR SGM Plus

6-FAM
Blue

D3S1358 vWA D16S359 D2S1338

JOE
Green

X-Y D8S1179 D21S11 D18S51

NED
Yellow

D19S433 THO1 FGA

ROX
Red

AmpFlSTR Indentifiler

6-FAM
Blue

D8S1179 D21S11 D7S820 CSF1PO

VIC
Green

D3S1358 THO1 D13S317 D16S359 D2S1338

NED
Yellow
PET
Red

D19S433 vWA TPOX D18S51

X-Y D5S818 FGA

LIZ
Orange

PowerPlex 16 System

FL
Blue

D3S1358 THO1 D21S11 D18S51 Penta E

JOE
Green

D5S818 D13S317

D7S820 D16S539 CSF1PO Penta D

Tamra
Yellow
CXR
Red

X-Y D8S1179 TPOX

vWA FGA

100 bp 200 bp 300 bp 400 bp 500 bp

Figure 6.3 PCR multiplexes use up to five different dyes to label PCR products. The allelic ranges
of three commonly used multiplexes, the AmpF STR SGM Plus ,AmpF STR Identifiler and
the PowerPlex 16 are shown. The use of multiple dyes allows the detection of the internal-lane
size standard (ROX TM in SGM Plus, LIZ TM in Identifiler and CXR in PowerPlex 16) and three or
four overlapping STR loci, where the use of different dyes allows the alleles to be assigned to the
correct locus