The protocol presented here provides an avenue to multiplex more than 300 markers in a single pool. The primer pools described therein have low marker complexity with approximately 12 markers per pool. Multiplexing on a small scale has been demonstrated by (Tamang et al. Genotyping by Multiplexed Sequencing (GMS) is a highly adaptable and effective genotyping tool for tracking markers of interest with high accuracy, which can translate to a more efficient and affordable MAS process in future breeding projects. Marker assisted selection (MAS) has been an effective tool for ensuring the transfer of quantitative trait loci (QTL) with high effects into elite barley lines. Marker-assisted selection provides opportunities for enhancing the response from selection because molecular markers can be applied in early generations, with high precision and reductions in cost (Miedaner and Korzun 2012). Here we describe a more efficient and adaptable protocol for tracking important barley traits for marker-assisted selection (MAS). Many barley traits contribute to malting quality, as well as food and feed quality. The United States produced approximately 169.6 million bushels of barley at an estimated price of 761.2 million dollars in 2019 (USDA NASS). Good markers have an adequate read depth of at least 5 amplicons and are reliably present across the population.īarley ranks a distant 4th in grain production worldwide with a total of 140.6 metric tons annually. We have identified 577 SNP markers that work well using this protocol providing adequate genome coverage for genomic selection and tag 267 QTL’s for genes of interest. In this study we have taken the GMS protocol developed in wheat and adapted it to barley. (2) The marker sets are easily customized to the species of interest and can readily be changed. The advantages of this protocol include: (1) This PCR-based direct sequencing protocol generates information from markers of interest and provides a more streamlined and accurate analysis process, by multiplexing hundreds of informative markers into a single sequencing run. The genotypes derived from direct sequence reads reduce ambiguity in genetic analysis. We have developed a robust protocol called genotyping by multiplexed sequencing (GMS) using SNP markers, providing informative genotypic data with greater flexibility. However, these genotyping technologies have some drawbacks that limit their usefulness in analysis. Genotyping by sequencing (GBS) and single nucleotide polymorphism (SNP) chip technologies are the primary SNP genotyping technologies used today.

Mtasa on marker hit size