The Thraupidae family is one of the most wanted by bird breeders in Brazil due to its diverse, colorful and melodious singers as representatives. The Great-billed Seed-finch, Sporophila maximiliani, is the only representative of the genus Sporophila considered critically endangered in Brazil. Due to the demands of environmental agencies and of conservation programs, there is a need to increase the number of molecular markers available for the genus and specially for S.maximiliani. Therefore, this work aimed to provide a new set of microsatellite markers for S. maximiliani in order to help bird breeders and environmental agencies on fulfilling its demands as well as contributing with extra genetics tools for conservation programs of the Great-billed Seed-finch. Of the 30 markers developed, 25 successfully amplified and 22 were polymorphic. Annealing temperature varied from 52 to 64°C, number of alleles from two to 13 and the medium allele richness was 7.25 and medium expected and observed heterozygosity was, respectively, 0.812 and 0.661. The identity estimate was 8.54x10 -27 and all the other probabilities of non-exclusion (sib-identity, parent pair and first-parent) were <0.001, indicating that this set of microsatellite markers have high genetic variability and high power of individual genetic differentiation for S.maximiliani. Therefore, this work increases the options of molecular markers to be used on inspection for environmental agencies and for conservation programs on analyzing genetic variability and population studies for the Great-billed Seed-finch.

Genomic DNA was extracted from blood samples for all of the analysis in this work through Qiagen DNeasy Blood & Tissue kit (QIAGEN, 2006). Contigs generated for the draft genome of S. maximiliani (GenBank accession number: MF327582) (Ludwig et al. 2017) were accessed to identify microsatellite regions using the software MSATCOMMANDER (Faircloth 2008). Over 100,000 contigs containing microsatellite regions were identified, and about 1,000 primer pairs suggested (Dryad: xxx). The primer pairs were screened for the generated fragment size (amplicons of 75–300 bp), and selected microsatellites containing at least seven repeats of tri/tetra-nucleotide motifs, because microsatellites with tri- and tetra-nucleotide motifs can be easily discerned, avoiding issues such as stuttering pattern (e.g. de Valk et al. 2005). Microsatellite markers were designed for 30 loci with motifs varying from four to six nucleotides.