Taxonomic and molecular characterization of 15 wild-growing tulip species of Greece using the internal transcribed spacer (ITS) nuclear marker in combination with the psbA-trnH and trnL/trnF plastid markers

Abstract

Due to the absence or scarcity of DNA sequence data in GenBank regarding Mediterranean, Balkan or Greek native tulip species, we performed molecular characterization of the 15 wild-growing Greek tulip species through analysis of the internal transcribed spacer nrDNA and the psbA/trnH and trnL/trnF cpDNA regions. To facilitate conservation and sustainable utilization efforts, the aim was to identify and complement the genetic distinctiveness of six Greek endemic and five Balkan or Aegean endemic and/or subendemic wild-growing Greek native tulips and two naturalized ones of Asiatic origin, among which seven are threatened with extinction. Molecular phylogenetic analysis of 148 DNA sequences was conducted, and 85 Tulipa taxa (species and subspecies) were aligned and compared to 40 new DNA sequences generated herein for the 15 wild-growing Greek tulips. These three molecular markers resulted in 354 single-nucleotide polymorphisms (SNPs) and 20 distinct indels for all Tulipa taxa, of which 100 SNPs and 12 indels were related to the Greek tulips. Phylogenetic dendrograms were constructed for all the above-mentioned molecular markers, and the DNA sequences were analyzed separately and jointly, leading to three distinct clusters for the 15 wild-growing Greek tulips. Group 1 included Greek members of subgenus Tulipa, that is, T. scardica, T. undulatifolia, T. rhodopea, T. agenensis and T. raddii; Group 2 included only T. clusiana (subgenus Clusianae) and Group 3 included Greek members of subgenus Eriostemones, that is, *T. bakeri, T. saxatilis, T. australis, *T. cretica, *T. goulimyi, *T. orphanidea T. bithynica, *T. hageri and *T. doerfleri (asterisks indicate single-country endemics).

Keywords

• Barcoding
• PCR
• botanical collections
• tulip bulbs
• SNPs
• indel