Short Tandem Repeats (STRs) are efficient tools for mapping specific traits or to follow the flow of genetic material in a population. The technology is based on the presence of short tracks of di, tri, tetra or penta nucleotide repeats which are common in the genomes of eukaryotic organisms. These short tracks (5-10 repeating units) are faithfully transmitted through sexual reproduction but are often highly polymorphic within a population.
As part of our continuing efforts to fully characterize and authenticate our client's cell lines, MDS offers genotyping of individual samples at STR loci using conventional techniques. STR loci are among the most informative polymorphic markers in the genome. The fingerprinting process used at MDS involves simultaneously amplifying eight STR loci (D5S818, D13S317, D7S820, D16S539, vWA, TH01, TPOX, CSF1PO) and the amelogenin gene in a multiplex PCR reaction which allows for discrimination of fewer than 1 in 108 individuals.
Genomic DNA is purified, and the short tandem repeat (STR) regions of interest are amplified using the polymerase chain reaction (PCR). The amplification products, generally less than 500bp in size, are analyzed on a denaturing polyacrylamide gel. Allelic ladders containing fragments of the same size as several or all known alleles are run on the gel alongside the amplified samples. Comparison of sample and allelic ladders allows easy interpretation of amplified alleles. After electrophoresis, alleles are detected by silver staining, fluorescent detection or the use of radioactivity.
Amplified fragment length polymorphisms can be divided into two categories: Long Tandem Repeats (LTRs) and Short Tandem Repeats (STRs). Both LTRs and STRs can be amplified using the polymerase chain reaction. One of the advantages of analyzing PCR-based variable number tandem repeat systems is that individual alleles can be resolved into more discrete fragments than with RFLP analysis. Discrete alleles enable phenotype determination by reference to allelic ladders in adjacent lanes of the gel or, in the case of some fluorescent-based detection systems, by reference to an internal lane standard