Variant Allele Sequencing


Participants: Margaret C. Kline and John M. Butler


Project Timeframe: May 2002 to present


Purpose: Variant alleles or "off-ladder alleles" exist as do polymorphisms in the flanking regions of STRs that can cause allele dropout when they fall underneath PCR primer binding sites. Characterization of the molecular basis for this genetic variation is beneficial to the forensic DNA community and other researchers who may want to avoid these polymorphisms when designing PCR primers for future assays.


Progress: For the past several years, our group at NIST has been sequencing unusual STR alleles provided by members of the human identity testing community. Some of these alleles possess point mutations in primer binding sites leading to allele dropout ("null" alleles) while others are "off-ladder" variants. Our strategy has been to develop new PCR primers that anneal outside all known STR kit primer positions for the common forensic STR loci. With this approach it is then possible to examine the nucleotide sequences underlying these primer binding sites when performing DNA sequence analysis.  Sequencing these unusual alleles helps to reveal the molecular basis for their variation. Summaries of our findings (along with who provided the sample) are listed on a separate STRBase section:  

If you wish to provide a sample for testing (cost is covered by our NIJ funding), we will need at least 10 ng of genomic DNA. Please contact either Margaret Kline (301-975-3134; or John Butler (301-975-4049; if you have questions about submitting a sample for STR allele sequencing.


Publications or Presentations Resulting From This Project:

Margaret Kline talk at 57th Annual Meeting of the American Academy of Forensic Sciences (New Orleans, LA), February 25, 2005, "Some Interesting Point Mutations and Deletions Found Through STR Allele Sequencing" [.pdf]


Margaret Kline talk at Bode East Coast Advanced DNA Technology Workshop (Duck Key, FL), May 24, 2005, "Finding Point Mutations, Deletions and New Alleles Through STR Allele Sequencing" [.pdf]


Barni, F., Salata, E., Rapone, C., Kline, M., Lago, G., Butler, J., Berti, A. (2007) Characterisation of novel alleles and duplication events in the STR DYS19, DYS439, DYS389II and DYS385 loci. Identity (Promega Newsletter for Europe, Middle East and Africa), Spring 2007, pp. 1-3. [.pdf]


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Last updated: 06/20/2007


Disclaimer: This project was supported by National Institute of Justice Grant Number 2003-IJ-R-029, which is an interagency agreement between NIJ and the NIST Office of Law Enforcement Standards, awarded by the National Institute of Justice, Office of Justice Programs, US Department of Justice. Points of view in this document are those of the authors and do not necessarily represent the official position or policies of the US Department of Justice. Certain commercial equipment, instruments and materials are identified in order to specify experimental procedures as completely as possible.  In no case does such identification imply a recommendation or endorsement by the National Institute of Standards and Technology nor does it imply that any of the materials, instruments or equipment identified are necessarily the best available for the purpose.