Metrological Support of Nucleic Acid Sequence Analysis

Abstract

Metrological support for the process of molecular genetic identification is proposed. An establishment of a nucleotide sequence is the main objective of the DNA/RNA analysis. Such nucleotide sequence provides key information about the role and function of nucleic acids in living systems, as well as the origin of biological objects during molecular genetic identification. The molecular genetic identification is based on the modern methods of studying DNA sequences, which are inherited by the living organisms from generation to generation. In some cases, comparison of DNA sequences is required to establish the authenticity of biological objects, as well as determine biological affiliation or kinship with other biological objects. The problems of metrological support of molecular genetic identification by Sanger DNA sequencing using fluorescent-labeled reaction terminators and capillary electrophoresis are considered. Methods and means of metrological support of the DNA/RNA analysis were developed, which include the determination of a nucleotide as a nucleic acid monomer, the use of a nucleotide sequence of nucleic acids used in certified methods and procedures, the use of a genetic analyzer as a measurement tool along with related auxiliary equipment, as well as the use of a standard sample of a fragment of human mitochondrial DNA has been developed and approved. A Nanofor 05 domestic genome analyzer was tested. The following two procedures were developed and certified: for measuring the nucleotide sequence of the section of the control region of mitochondrial DNA of fish (sturgeon and paddlefish families) by Sanger sequencing, and for measuring the nucleotide sequence of the 5' region of the COI gene of the mitochondrial DNA of aquatic biological resources by Sanger sequencing. Both measurement procedures are based on the use of fluorescent-labeled reaction terminators and capillary electrophoresis. The results are important for use in the DNA studies in the field of biological sciences, food industry, biotechnology, and DNA forensics.