Development of Inter-Retrotransposon Amplified Polymorphism (IRAP) Markers and DNA Fingerprinting of Blueberry Accessions

Blueberries (Vaccinium spp.) are valued for their nutritional benefits but face challenges in germplasm identification, phylogenetic analysis, and breeding due to their complex genetic background. Long Terminal Repeat Retrotransposons (LTR-RTs), major drivers of plant genetic diversity, offer a basis for the Inter-Retrotransposon Amplified Polymorphism (IRAP) system, which excels in germplasm identification, diversity assessment, and relatedness studies. Here, we developed a blueberry IRAP system using Ty1-copia reverse transcriptase sequences. From 25 core primers, we obtained 266 polymorphic loci (average PIC = 0.866). These IRAP markers fingerprinted 112 accessions and revealed relationships through Nei’s diversity index (H = 0.361), Shannon’s index (I = 0.533), AMOVA (9.33% among regions; 90.67% within populations; Nm = 1.50), UPGMA dendrograms (three clusters at 0.615 similarity), and PCoA, indicating weak geographic structure across Guiyang, Qiandongnan, and Bijie consistent with Nm = 1.50 (homogenizing gene flow). The dendrogram and PCoA indicate among-accession heterogeneity with weak geographic structuring across Guiyang, Qiandongnan, and Bijie, consistent with the AMOVA and gene-flow estimates. We also built a Molecular IDs database for differentiation. IRAP proved highly efficient for identification and analysis, matching SSR/SNP polymorphism levels while offering advantages like low-cost agarose gel resolution for detecting subtle clonal variants in polyploids—outperforming SSRs in field triage and complementing SNPs’ high throughput. This supports IP protection and breeding. Together with established SSR/SNP platforms, this IRAP approach can support IP protection and breeding as a complementary, cost-effective option.