Allelic ladders and reference genotypes for a rigorous standardization of poplar microsatellite data

Abstract

A correct identification of members of the poplar hybrid complex Populus × canadensis is essential in breeding programs and studies in introgressive gene flow. Molecular marker protocols have been developed for this purpose. However, due to missing standards, these techniques have so far not been suited to the transfer of results between different laboratories. We present here a powerful system of nuclear microsatellite DNA (nSSR) fingerprints, standardized by allelic ladders and reference genotypes. Seven nSSR loci provided fingerprints of 65 commercial poplar clones. Their alleles were used to construct allelic ladders. Thus, a first standardized register of poplar clones is now available. All procedures were optimized according to simplified DNA extraction protocols, multiplexed PCR and electrophoresis procedures. Corresponding data originating from two different electrophoretic platforms in different laboratories were congruent when the allelic ladder was used. Unambiguous differentiation of the clones was based on a very low probability of identity (PI) of 1.95 × 10−8. Our results revealed discrepancies between clone denotations and genetic fingerprints. This suggests that, potentially, members of the clone collection could have been mixed up, thus confirming the demand for rigorous standards. The protocol presented can be exploited in a manifold way, e.g. to enlarge the present clonal molecular data base, or to use it for purposes of certification and control. Furthermore, the allelic ladders are recommended for use in poplar population genetic studies across different laboratories. The allelic ladders and single sample reference genotypes can be obtained on demand.