Validating Blackberry Seedling Pedigrees and Developing an Improved Multiplexed Microsatellite Fingerprinting Set

Abstract

Confirming parentage and clonal identity is an important aspect of breeding and managing germplasm collections of clonally propagated, outcrossing crops, like blackberry (Rubus subgenus Rubus). DNA fingerprinting sets are used to identify off-cross progeny and confirm clonal identity. Previously, a six-simple sequence repeat (6-SSR) fingerprinting set was developed for blackberry using a small number of samples. The usefulness of the 6-SSR fingerprinting set for pedigree confirmation had not been evaluated. Therefore, it was used in this study to validate parentage for 6 and 12 biparental populations from the University of Arkansas (UA) and US Department of Agriculture Agricultural Research Service (USDA-ARS), Horticultural Crops Research Unit (HCRU) breeding programs, respectively. Twenty-seven of the 489 individuals in these breeding populations were identified as off-cross. The 6-SSR fingerprinting set was sufficient for parentage confirmation; however, a total of 61 plants distributed across 28 sets of genotypes could not be distinguished from each other. An 8-SSR fingerprinting set with improved resolution was subsequently developed and used to evaluate 177 Rubus accessions from the USDA-ARS National Clonal Germplasm Repository, UA, and USDA-ARS HCRU programs. The 8-SSR fingerprinting set distinguished all samples expected to have unique genotypes and identified differing DNA fingerprints for two sets of accessions suspected to have identical fingerprints. Cluster analysis grouped the accessions from the eastern and western US breeding programs based on geography and descent. Future work will focus on establishing a database of DNA fingerprints for germplasm identification and for determining pedigree relationships between blackberry accessions.