Development of Multiplexed Marker Sets to Identify the Most Relevant Poplar Species for Breeding

Abstract

Within the genus Populus, about 30 species are classified into six sections, of which some are cross-compatible. Besides naturally occurring hybrids, huge breeding programs have led to a high number of artificially produced hybrids, for which the determination of genetically involved species by morphological characteristics is often difficult. This necessitates the use of molecular markers for the identification of both maternal as well as paternal species, and in the case of complex hybrids, the genealogy. For this reason, we developed new chloroplast and nuclear markers for the differentiation of up to 19 poplar species, with one to 32 individuals per species regularly used in breeding programs based on already known barcoding, other chloroplast regions, and nuclear genes of interest. We developed methods to identify species by either species-specific nucleotide variations or, when no initial information for the species was given, by using a set of markers either in a procedure of exclusion or in a multiplexed marker set. The developed markers can all be used with low-cost equipment, and some can additionally be applied using a genetic analyzer. We combined these markers in multiplexes for a very fast and easy-to-use application for the identification of poplar species and their hybrids.

Development of 5S rDNA-Based Molecular Markers for the Identification of Populus deltoides Bartr. ex Marshall, Populus nigra L., and Their Hybrids

“…In cases where the polymorphic regions are short and cannot serve as a basis for primer development, molecular markers of other types can be created. For example, short polymorphic regions can be used to develop RLFP, SNP, or CAPS (cleaved amplified polymorphic sequences) markers [14,27]. Thus, the approaches for the development of species-specific markers presented in the article are not universal.…”

“…Attempts to use DNA barcoding for Populus species identification have been made worldwide, but a single polymorphic locus (single nucleotide polymorphism, SNP) for all poplars has not been determined. Therefore, systems of different markers for poplar barcoding (and PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) markers based on these SNPs) are repeatedly offered by many researchers [11][12][13][14].…”

Application of retrotransposon-based Inter-SINE Amplified Polymorphism (ISAP) markers for the differentiation of common poplar genotypes

“…Many different types of nuclear markers have been tested and established to differentiate and identify poplars at genus, species or clone level. For example, single nucleotide polymorphisms (SNPs) have been developed that are suitable for species discrimination in poplar (Schroeder et al 2017). However, this requires the sequencing of amplified fragments in order to identify genotype-specific base substitutions, which makes this method complex and time-consuming.…”

The Development of Populus alba L. and Populus tremula L. Species Specific Molecular Markers Based on 5S rDNA Non-Transcribed Spacer Polymorphism

“…Thus, the development of biochemical and DNA molecular markers has been actively applied to work with poplar species and their hybrids [16,17]. Different types of DNA markers were previously developed, though microsatellite and single nucleotide polymorphism (SNP) markers are the most commonly used [18][19][20][21]. Also, 5S rDNA non-transcribed spacer (NTS) based markers (sequence-characterized amplified region (SCAR) markers) have been recently designed for the identification of some poplars (P. nigra, P. deltoides, and P. × canadensis) in the Aigeiros section [22].…”