Rapid screening for Native American mitochondrial and Y-chromosome haplogroups detection in routine DNA analysis.
Aiming to detect individuals of Native American maternal or paternal ancestry a rapid screening approach has been developed. Its strategy was based on SNP typing by Real Time PCR (rt-PCR) followed by High Resolution Melting analysis (HRM). After extraction, DNA was quantitated by rt-PCR using commercial kits; samples were then submitted to two multiplex reactions in order to determine the major Native American mtDNA and Y-chromosome haplogroups by HRM. One cocktail included primers flanking nucleotide substitutions that define mtDNA haplogroup C and sub-haplogroups A2, B2, and D1. The other included primers flanking Y-SNPs M3, M269 and U179 that allowed discriminating Q and non-Q haplogroups. In all cases amplicons were <125 nucleotides long in order to increase the peak resolution. The accuracy of the results obtained was established by means of sequencing analysis of the amplicons. The new working-flow here proposed facilitates and speeds-up the screening process that may preclude a detailed sequencing analysis of particular samples, or for further molecular epidemiological investigations in which continental origin influences might be relevant.
Zuccarelli
Gala
G
Servicio de Huellas Digitales Genéticas y Cátedra de Genética y Biología Molecular, School of Pharmacy and Biochemistry, Buenos Aires University, Argentina.
Alechine
Evguenia
E
Caputo
Mariela
M
Bobillo
Cecilia
C
Corach
Daniel
D
Sala
Andrea
A
D001118:Argentina; D041322:Chromosomes, Human, Y; D016172:DNA Fingerprinting; D004272:DNA, Mitochondrial / Q000235:genetics*; D005006:Ethnicity / Q000235:genetics*; D005828:Genetics, Population; D006239:Haplotypes; D006801:Humans; D008297:Male; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D017421:Sequence Analysis
D004272:DNA, Mitochondrial