Standards for Mitochondrial Research and Mutation Detection

Current research focuses primarily on the use of human mitochondrial DNA for disease diagnostics. We have developed two Standard Reference Materials (SRMs) for the amplification and sequencing of the entire human mitochondrial DNA, and one SRM for the detection of low-frequency mutations or single nucleotide polymorphisms (SNPs) in either mtDNA or in pooled nuclear DNA, or in heteroplasmic sites in mtDNA. Mitochondrial mutations also play a role in human identification (forensics).

• SRM 2392 “Mitochondrial DNA Sequencing (Human)” ( https://srmors.nist.gov/view_cert.cfm?srm=2392 ) is intended to provide quality control when performing the polymerase chain reaction (PCR) and sequencing of human mitochondrial DNA for forensic identifications, medical diagnosis, or mutation detection. This SRM is composed of well-characterized extracted human DNA from CHR and GM09947A and cloned DNA from the HV1 region of CHR. .
• SRM 2392-I contains the HL-60 template recommended by the FBI for use as a positive control for forensic laboratories and can be used in conjunction with SRM 2392. (https://srmors.nist.gov/view_detail.cfm?srm=2392-I)
• SRM 2394, “Heteroplasmic Mitochondrial DNA Mutation Detection Standard”( https://srmors.nist.gov/view_cert.cfm?srm=2394 ) is composed of human mitochondrial DNA mixtures which simulate different levels of heteroplasmy or low-frequency mutations in pooled samples. SRM 2394 is intended to provide quality control benchmarks for forensic, medical, and DNA scientists to assess the detection sensitivity of low-frequency mutations, single nucleotide polymorphisms (SNPs) in either mitochondrial DNA or in pooled nuclear DNA samples, or heteroplasmic sites in mtDNA.
• An interactive web site, http://www.cstl.nist.gov/biotech/strbase/mitoanalyzer.html has been developed to allow investigators to search for information on the mitochondrial mutations found in their samples: specifically, which gene is affected, whether it causes an amino acid change, the nature of that change, the position in the protein, and the amino acid sequence of the new protein, and if the change is associated with a disease.

• B. C. Levin, K. L. Richie, M. C. Kline, J. W. Redman, D. K. Hancock, “Human DNA Standard Reference Materials Developed by the National Institute of Standards and Technology, ” in “Progress in Genome Research” , Nova Science Publishers, in press.
• D. K. Hancock, L. A. Tully, B. C. Levin, “A Standard Reference Material to determine the sensitivity of techniques for detecting low-frequency mutations, SNPS and heteroplasmies in mitochondrial DNA,” Genomics, 2005, accepted.
• B.C. Levin, K. A. Holland, D. K. Hancock, M. Coble, T. J. Parsons, L. J. Kienker, D. W. Williams, M. P. Jones, K. L. Richie, “Comparison of the Complete mtDNA Genome Sequences of Human Cell Lines - HL-60 and GM10742A - from Individuals with Pro-myelocytic Leukemia and Leber Hereditary Optic Neuropathy, Respectively, and the Inclusion of HL-60 in the NIST Human Mitochondrial DNA Standard Reference Material - SRM 2392-I,” Mitochondrion 2003, 2, 387 - 400.
• B. C. Levin, D. K. Hancock, K. A. Holland, H. Cheng, K. L. Richie, “Human Mitochondrial DNA - Amplification and Sequencing Standard Reference Materials – SRM 2392 and SRM 2392-I ,' NIST Special Publication 260-155 (2003).
• D. K. Hancock, F. P. Schwarz, F. Song, J. J. C. Wong, B. C. Levin, “Design and Use of a Peptide Nucleic Acid for Detection of the Heteroplasmic Low-Frequency Mitochondrial Encephalomyopathy, lactic Acidosis, and Stroke-like Episodes (MELAS) Mutation in Human Mitochondrial DNA,” Clin. Chem. 2002, 48, 2155-2163.