Genetic variation of melon (C. melo) compared to an extinct landrace from the Middle Ages (Hungary). I. rDNA, SSR and SNP analysis of 47 cultivars

Microsatellite profiles of 47 melon cultivars and landraces were analyzed and compared to the aDNA (ancient DNA) of seed remains from an extinct sample recovered from the 15th century (Budapest, Hungary). An aseptic incubation followed by ITS (internal transcribed spacer) analysis was used to exclude the exogenously and endogenously contaminated medieval seeds and to detect SNPs (single nucleotide polymorphism) in ITS1-5.8S-ITS2 region of rDNA (ribosomal DNA). SNPs were observed at the 94–95 bp (GC to either RC, RS or AG) of ITS1; and at 414 bp (A-to-T substitution), 470 bp (T to Y or C), 610 bp (A to R or G) and 633 bp (A-to-G transition) of ITS2. For comparative microsatellite analysis SSRs (simple sequence repeats) detected by ALF (automated laser fluorometer) was used. Eight of the 20 SSR primer pairs amplified 40 microsatellite alleles in identical fragment ranges. A total of 485 alleles were detected in the 47 melon cultivars. The number of alleles per marker ranged from 2 to 7 with an average of 5.7 including CMCT44 (2 alleles), CMAG59 (5 alleles), CMGA104 (5 alleles), CMCT134 (4 alleles), CMTA134 (6 alleles), CMCTT144 (7 alleles), CMTC168 (6 alleles) and CMCT170 (5 alleles). Sequence analysis of the microsatellite alleles showed different fragment lengths depending on changes in the number of unit of core sequences. Dendrogram produced by SPSS11 based on the presence versus absence of SSR alleles revealed that medieval melon had the closest genetic similarity to a registered melon cultivar Hógolyó selected from an old Hungarian melon landrace. These results also indicated that cloned DNA sequences recovered from aDNA of medieval melon can be of use for molecular breeding of modern melon cultivars via gene transfer