The Common Pheasant (Phasianus colchicus) Linnaeus, 1758 is the most widespread pheasant in the world with a natural, geographic range spanning in temperate to subtropical regions of the Palearctic realm (Johnsgard 1999). This species exhibits a high-level of intra-specific differentiation in plumage coloration and patterns in males. Thirty subspecies forming five subspecies groups were defined mainly based on geographically distributed affinities and morphological characters (Cramp and Simmons 1980; Johnsgard 1999; Madge and McGowan 2002). The five subspecies groups are as follows: (1) the colchicus group (Black-necked Pheasants, west and south of the Caspian Sea, including the subspecies persicus, talischensis, colchicus and septentrionalis); (2) the principalis-chrysomelas group (White-winged Pheasants in Central Asia, including the subspecies principalis, zarudnyi, chrysomelas, bianchii, zerafschanicus and shawii); (3) the tarimensis group (Tarim Pheasant, tarimensis in Tarim Basin in southeastern Xinjiang, China); (4) the mongolicus group (Kirghiz Pheasants in northern Xinjiang, China and eastern Kazakhstan, comprising mongolicus and turcestanicus) and (5) the most subspecies-rich group, the torquatus group (Grey-rumped Pheasants, mostly found in China, containing 17 subspecies: decollatus, satscheuensis, pallasi, suehschanensis, torquatus, kiangsuensis, rothschildi, karpowi, strauchi, elegans, vlangalii, hagenbecki, edzinensis, alaschanicus, sohokhotensis, takatsukasae and formosanus) (Madge and McGowan 2002).
The Common Pheasant has a long history of captivity and being introduced as a common game species in western Europe, North America and Australia (Hill and Robertson 1988; Johnsgard 1999). This species deserves conservation management and sustainable use for several reasons. First of all, because natural populations of the Common Pheasant have been dramatically declining due to the loss of its natural habitats, hunting and other anthropogenic disturbances (Sotherton 1998), restocking of this bird is increasingly needed. For example, native habitat loss due to reclamation for agriculture caused a population decline in the subspecies principalis and persiscus in Iran (Solokha 1994). The subspecies turcestanicus is probably extinct now as a result of the aridification of the Aral Sea (Lepage 2007). As well, hybridized decendents between local subspecies and ex situ subspecies, due to artificial introduction of captive birds for hunting purposes, are evident in the wild (Braasch et al. 2011). Even worse is likely to occur, in so far as Common Pheasants in the wild may interbreed with a commercial, captive breed, the so-called “seven-color wild pheasant” which is a hybridized race between the Common Pheasant and its sister species, endemic to the Japan archipelagos, the Green Pheasant (Phasianus versicolor). For all these reasons genetic pollution in the wild Common Pheasant gene pool may occur. Last but not least, some range-restricted subspecies of the Common Pheasants inhabit isolated range and extreme environments such as arid regions, islands and mountains which preserve unique phenotypes and genotypes for future conservation and stocking (Braasch et al. 2011; Kayvanfar et al. 2017). For example, the formosanus subspecies of the Common Pheasant is endemic to Taiwan; other subspecies hagenbecki, alaschanicus and tarimensis are isolated and have adapted to semi-desert conditions (Johnsgard 1999). These conservation and management issues require evaluation using conservation approaches in genetics. Developing permanent genetic resources, such as autosomal microsatellites are of critical importance.
Microsatellites, also known as simple sequence repeats (SSRs), are a preferred type of markers in conservation genetics (Sunnucks 2000). Because of their heritable mode, SSRs usually have a higher mutation rate than that of mitochondrial and nuclear intronic markers and represent a very useful tool to genotype individuals and thus allow the quantifications of intraspecific genetic diversity, population structure and gene flow (Selkoe and Toonen 2006). Applications of SSRs are also reliable because of their relatively great abundance in genomes, high level of genetic polymorphism, co-dominant inheritance mode, analytical simplicity and repeatability of results across laboratories. So far, no species-specific microsatellites are available for the Common Pheasant although a previous study showed that cross-amplification of a very limited number of SSRs from other closely related Phasianinae species should be applicable for Common Pheasants (Baratti et al. 2001).
Recent advances in next generation sequencing (NGS) technologies enable the generation of large number of sequences efficiently and cost-effectively (reviewed in Ekblom and Galindo 2011). In addition, the so-called “Restriction-site Associated DNA” (RAD) method was consequently developed as a reliable means for genome complexity reduction (Baird et al. 2008). The concept is based on acquiring the sequence adjacent to a set of particular restriction enzyme recognition sites and then obtain sequences (RAD-seq) by NGS technology. Application of the RAD method, using the Illumina platform, has the advantage that it generates relatively long paired-end sequencing reads (100–150 bp), is cost-effective and sufficient to develop SSRs (Castoe et al. 2012). Another advantage is that a RAD-seq does not require a reference genome to be available and allows de novo assembly (Willing et al. 2011).
In this study we developed a set of autosomal microsatellites for the Common Pheasant using RAD-seq. We further designed multiplex PCR sets and tested for genetic polymorphism in a selected panel of 10 selected SSRs, which provide a tool for conservation genetic and studies of the evolution in the Common Pheasant.