Integrated tool for microsatellite isolation and validation from the reference genome and their application in the study of breeding turnover in an endangered avian population

Accurate individual identification is required to estimate survival rates in avian populations. For endangered species, non-invasive methods of obtaining individual identification, such as using molted feathers as a source of DNA for microsatellite markers, are preferred because of less disturbance, easy sample preparation and high efficiency. With the availability of many avian genomes, a few pipelines isolating genome-wide microsatellites have been published, but it is still a challenge to isolate microsatellites from the reference genome efficiently. Here, we have developed an integrated tool comprising a bioinformatic pipeline and experimental procedures for microsatellite isolation and validation based on the reference genome. We have identified over 95 000 microsatellite loci and established a system comprising 10 highly polymorphic markers (PIC value: 0.49-0.93, mean: 0.79) for an endangered species, saker falcon (Falco cherrug). These markers (except 1) were successfully amplified in 126 molted feathers, exhibiting high amplification success rates (83.9-99.7%), high quality index (0.90-0.97) and low allelic dropout rates (1-9.5%). To further assess the efficiency of this marker system in a population study, we identified individual sakers using these molted feathers (adult) and 146 plucked feathers (offspring). The use of parent and offspring samples enabled us to infer the genotype of missing samples (N = 28), and all adult genotypes were used to ascertain that breeding turnover is a useful proxy for survival estimation in sakers. Our study presents a cost-effective tool for microsatellite isolation based on publicly available reference genomes and demonstrates the power of this tool in estimating key parameters of avian population dynamics.