Sequence analysis of two de novo mutation alleles at the Y-STR locus

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• Abstract
• 1. Introduction
• 2. Materials and methods
• 3. Results and discussion
• Reference
• Copyright

Abstract 

We have performed haplotyping of 16 Y-STRs (giving 17 PCR products): DYS19, DYS389I, DYS389II, DYS390, DYS391, DYS392, DYS393, DYS385, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635, and Y GATA H4 in 47 biological paternity cases (father and son) in Japan. We have detected two unusual alleles at the DYS439 and DYS448 locus in two paternity trio cases. In one case, DYS439 allele 15 of the son was found not to derive from the father (allele 14). In the other case, the father (allele 18) and the son (allele 17) shared no bands at DYS448. Sequencing showed that the son gained a single unit (GATA) at DYS439 in one case and the son losed 1 unit (AGAGAT) at DYS448 in the other case.

Keywords: Short tandem repeat (STR), Sequence, DYS439, DYS448, Paternity testing, Mutation

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1. Introduction 

Short tandem repeats (STRs) are useful for personal identification and paternity testing because the number of repeats within STRs tends to be highly variable and these STR polymorphisms can be rapidly analyzed using PCR. Autosomal STR makers are widely applied to personal identification and paternity testing. However, in some cases of a brother, son, and uncle, from his paternal lineagetesting the investigation of the Y chromosomal STR markers may be more informative than the autosomal STRs [1]. We encountered two paternity cases where unusual alleles at the DYS439 and DYS 448 loci were detected in the son. In this study we characterized the mutant alleles at the DYS439 and DYS 448 loci by sequence analysis.

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2. Materials and methods 

Peripheral blood samples (1ml each) were obtained from paternity trios. Genomic DNAs were recovered from buffy coat by proteinase K digestion followed by sodium iodide extraction. Y chromosomal loci were amplified by Amp FℓY file typing kit (Applid Biosystems, Foster City, CA). PCR products of 1.5μl were diluted with 24.5μl of Hi-Di Formamide (Applied Biosystems), denatured at 95°C for 2min, and electrophoresed on POP4 by using ABI PRISM 310 Genetic Analyzer (Applied Biosystems). DYS439 and DYS448 alleles were sequenced on an ABI PRISM 310 Genetic Analyzer (Applied Biosystems) by using the Big Dye Terminator Cycle Sequencing FS Ready Reaction kit (Applied Biosystems).

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3. Results and discussion 

The repeat region of DYS439 consists of simple sequence (GATA). Sequencing showed that the son gained a single unit (GATA) at DYS439 in case 1. Allele 15 of the son seemed to result from gain of a single unit from allele 14 of the father in this case.

On the other hand, DYS448 was more complex in structure since this contained two variable blocks interspersed with intervening non-variant repeat blocks between the variants regions that were not counted in the repeat length nomenclature. The tandem repeat region could be subdivided into three different repeat types of 6bp length: whereas the first and third repeat ‘AGAGAT’ were variable forming the basis, the second repeat ‘AGAGAT’ was not polymorphic in all samples studied and was always present three times. Direct sequencing showed that the son lost 1 unit (AGAGAT) at DYS448 in case 2. In this case, allele 17 of the son must mutate from the father's allele 18 by loss of a single repeat unit.

In conclusion, STR markers used for paternity testing sometimes show unusual alleles due to mutation. It is necessary to sequence of these unusual alleles.

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Reference 

1. Jobling MA, Pandya A, Tyler-Smith C. The Y chromosome in forensic analysis and paternity testing. Int. J. Legal Med. 1997;110:118–124
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