Fast and non-invasive PCR sexing of primates: apes, Old World monkeys, New World monkeys and Strepsirrhines

Palle Villesen,Tina Fredsted

doi:10.1186/1472-6785-6-8

Abstract

BackgroundOne of the key tools for determining the social structure of wild and endangered primates is the ability to sex DNA from small amounts of non-invasive samples that are likely to include highly degraded DNA. Traditional markers for molecular sex determination of primates are developed on the basis of the human sequence and are often non-functional in distantly related primate species. Hence, it is highly desirable to develop markers that simultaneously detect Y- and X-chromosome specific sequences and also work across many species.ResultsA novel method for sex identification in primates is described using a triple primer PCR reaction and agarose gel electrophoresis of the sex-chromosomal isoforms of the ubiquitously transcribed tetratricopeptide repeat protein gene (UTX/UTY). By comparing genomic data from several mammals we identified the UTX/UTY locus as the best candidate for a universal primate sexing marker. Using data from several species we identified a XY-conserved region, a Y conserved region and an X conserved region. This enabled the design of a triple primer PCR setup that amplifies X and Y products of different length in a single PCR reaction.ConclusionThis simple PCR amplification of X and Y fragments is useful for sexing DNA samples from all species of primates. Furthermore, since the amplified fragments are very short the method can be applied to fragmented DNA extracted from non-invasive samples.

Highlights

One of the key tools for determining the social structure of wild and endangered primates is the ability to sex DNA from small amounts of non-invasive samples that are likely to include highly degraded DNA
The best hit was 248 bp with 95.56% identity between mouse and human. This region is a part of the Ychromosomal isoform of the ubiquitously transcribed tetratricopeptide repeat protein gene (UTY)
The UTY and the X-chromosomal homolog (UTX)/UTY region is suitable for triple primer design A small region in the alignment of 19 primate UTX/UTY sequences showed three distinct patterns, useful for primer design: The UTX/UTY primer is located in a region with high conservation between primate X and Y homologs

Summary

Introduction

One of the key tools for determining the social structure of wild and endangered primates is the ability to sex DNA from small amounts of non-invasive samples that are likely to include highly degraded DNA. The optimal marker would work on small amounts of non-invasive samples that are likely to include highly degraded DNA and be applicable in many species. In order to detect the male sex, a Y chromosomal fragment must be amplified This can be done by (a) amplification of a homolog region on X and Y with a length difference, (b) a triple primer PCR with a common primer X/Y and a Y specific primer (short fragment) and X specific primer (longer fragment) or (c) a multiplex PCR with a Y region and an positive control (autosomal or X region)

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