Systemic lupus erythematosus and rheumatoid arthritis

particular SNP, are mixed with forward and reverse primers, necessary reagents 
for the polymerase chain reaction (PCR) and the individuals genomic DNA. 
Each probe corresponds to one allele of the specific SNP and has a specific 
fluorophore and also a quencher, which absorbs the energy from the fluorophore 
when in close proximity. A short part of the template, which includes the SNP, 
is amplified with specific primers and if the right TaqMan probe is bound to 
sequence it will be a perfect binding and the DNA polymerase will digest the 
probe during the next cycle of polymerisation. This will separate the quencher 
from the fluorophore yielding a fluorescence that can be detected following 
exposure to a laser source. However, if the wrong TaqMan probe is bound there 
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will be a mismatch in binding and the whole probe will dissociate from the 
sequence upon polymerisation and no fluorescence can occur because of the 
proximity of the quencher. Any emitted fluorescence is detected by the 
dedicated TaqMan instrument and the results viewed as individual genotypes 
based on the intensity of the fluorescence (Figure 4). 
Figure 4.
Allelic discrimination of the 
PTPN22
1858C/T polymorphism in patients with early 
diagnosed RA. The x-axis denotes the intensity of fluorescence detected from the fluorophore 
specific for the probe that bound the T allele of the 
PTPN22
1858C/T polymorphism with a 
perfect match. The y-axis denotes the intensity from the fluorophore on the probe corresponding 
to the C allele. 
Based on the allelic discrimination plot in figure 4, there are two individuals 
who were homozygous for the T allele (TT) (red), indicating that the T allele 
was present on both chromosomes. In green are heterozygous individuals who 
have the C allele on one chromosome and the T allele on the other (CT). In blue 
are individuals homozygous for the C allele (CC) and in grey are the negative 
controls. If an allele is significantly more frequent in patients compared with 
controls, the allele is considered to be associated with the disease.
Association studies in case-control materials are now the commonest way of 
identifying candidate genes and causative alleles for complex diseases. There 
are two approaches when performing association studies: firstly, to investigate 
one or a few SNPs in a single candidate gene and compare the allele frequencies 
between cases and controls, and secondly, to perform a genome-wide 
association study (GWAS) comprising as many as one million SNPs across the 
genome. The latter is an ideal approach for identifying novel candidate genes. 
Both strategies require well-characterized case-control materials with patients 
and controls originating from the same geographical area. Population 
stratification might be a problem since allele frequencies may differ between 
individuals from different countries and from different ethnic backgrounds. In 
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unmatched case-control studies, population stratification could cause false 
positive/negative results.