The polymorphic markers of nuclear SSR (simple sequence repeat), SNP (single nucleotide polymorphism)/ InDel (insertion–deletion length polymorphism), and cpSSR (chloroplast SSR) were used to evaluate the genetic diversity of 52 trifoliate oranges and their relatives. The two genomic DNA-based markers produced high values of PIC (polymorphic information content) at 0.45 for SNP/InDel and 0.32 for nuclear SSR, indicating that there is a large diversity of genetic variationvwithin the trifoliate oranges. Three UPGMA (unweighted pair-group method with arithmetic means) dendrograms were generated based on the results of three marker systems. At 0.76 of coefficient, all of Poncirus accessions were clustered into one group, except the 74-1 Early Poncirus, one of the three precocious trifoliate oranges, which was placed separately into another group by nuclear SSR. At a higher coefficient, the trifoliate oranges were discriminated into 4, 6 and 6 groups by nuclear SSR (0.86), SNP/InDel (0.95), and cpSSR (0.95), respectively. The results revealed that three precocious trifoliate oranges were originated from different sources. Hubei Early Poncirus and Beibei Early Poncirus were the asexual mutants of different trifoliate oranges, and 74-1 was probably a sexual hybrid. The present study also indicated that ‘Zhiwenzhou,’ a diploid chimera of trifoliate orange and Satsuma is more closely related to Satsuma rather than to trifoliate orange, which is in concordance with the results of morphological observation.
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