Single-Nucleotide Polymorphisms (SNPs)

Single Nucleotide Polymorphisms (SNPs) for Human Identification An Emerging Forensic Tool

Genetic Typing is a powerful forensic tool used for investigating criminal and paternity cases. Current methods for DNA profiling reduces sample consumption, multiple manipulation procedures, reduce the processing time, and also amplifies 16 different loci through multiplexing. DNA Typing increases the power of discrimination in the identification of profiles. Current methods are Polymerase Chain Reaction and Short Tandem Repeats (STR) Typing.  Short Tandem Repeats have smaller product sizes, and the polymorphic nature of STRs makes them amenable to automation. Some of the samples are not amenable to STR typing due to low concentration of DNA samples or degradation. In order to overcome this issue, human mitochondrial DNA can be sequenced. Mitochondrial genome sequencing service is high, and only a few labs offer this service. The power of discrimination is very low in Mitochondrial DNA Typing.

Single Nucleotide Polymorphisms


One of the promising classes of genetic markers to type degraded or complex DNA samples containing a low level of DNA is Single Nucleotide Polymorphism (SNP). SNPs are base substitutions, insertions, or deletions that occur at a single position within the human genome. SNPs are abundant and occur every 100-300 base pairs along the DNA strand. The smaller product size is suitable for degraded DNA samples and can provide specific Identity about age, lineage, Identity, ancestry, and phenotype.

Single Nucleotide Polymorphisms are not represented in the CODIS database, whereas STR loci are very well established. SNP significantly reduces the cost and enhances abilities to resolves samples containing more than one contributor. SNP does not replace STR typing, but it will be an additional tool in DNA Typing. There are several applications for SNP- Hybridization based method and Enzyme based method. SNP genotyping is commercially available in the SNapshot multiplex system. The reaction tube observes 10 SNPs at specific sites along with the DNA in the Snapshot multiplex system. The Snapshot multiplex system is a primer-based extension assay consists of a dideoxynucleotide (ddNTP) single-base extension of an unlabeled nuclear primer. Each primer binds to a complementary strand upstream of an SNP nucleotide, and DNA polymerase extends primer by one fluorescently labeled ddNTP to SNP nucleotide. Since ddNTP is missing the 3’OH end, no more dNTP can be added. The incorporated nucleotides can be detected using electrophoresis in ABI Genetic Analyzer and allow SNP alleles to be determined using Genemapper software.

Single Nucleotide Polymorphisms


Novel SNPs can be detected using the LI-COR infrared DNA analyzer in human mitochondrial DNA and also in inbred strains. In one of the research studies, two segments in the coding region of mitochondrial DNA (NADH dehydrogenase subunit 2 genes)  and also hypervariable region was selected. These regions were sequenced, after amplification using forward primers and LI-COR’s IRDye™ labeled dideoxynucleotides.  No unique SNPs saw in the hypervariable region. Two SNPs were seen in the dehydrogenase gene and appeared to be of Asian descent. In the other study, researchers used  Smits et al. used the enzymatic cleavage of heteroduplexes using the endonuclease CEL I for SNP discovery. DNA was isolated from cell samples andwere subjected to PCR. The heteroduplexes cleaved with the endonuclease CEL I at the mismatched sites. The purified products were separated and sequenced on the 4300 DNA Genetic Analyzer.

Single Nucleotide Polymorphisms

DNA is extracted from both questioned and reference samples and placed in tubes or 96-well plates for screening.  During amplification, primers are labeled with a specific IRDye. After amplification, the heteroduplexes are cut at the mismatch sites by CEL I. (c) The cut strands are separated by electrophoresis in a genetic analyzer. SNPs in one fluorescence channel (IRDye 700) are confirmed by the presence of the opposite strand in the same lane in the other fluorescence channel (IRDye 800). Empty lines indicate no genetic variation or SNPs present in the sample. SNP discovery using endonuclease CEL I.

There are four different categories of Single Nucleotide Polymorphisms that benefit the forensic community


  • Identity testing- This test identifies an individual based on evidentiary testing and mixtures and degraded DNA samples.
  • Ancestry Informative SNPs- These SNPs give a high probability of an individual’s ancestry being from one part of the world or derived from two or more areas of the world.
  • Phenotypic Informative SNPs- Phenotypic SNPs provide a high probability of predicting phenotype.
  • Lineage Informative SNPs- Lineage informative SNPs identify relatives with high probabilities than simple biallelic SNPs.

The purpose of this section provides an overview of Single Nucleotide polymorphism and the use in the Forensic community.

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