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Background    Lilium  is an important ornamental bulb, possesses medicinal properties, and is also edible. Species within the  Lilium  genus share very similar morphology and macroscopic characteristics, thus they cannot be easily and clearly distinguished from one another. To date, no efficient species-specific markers have been developed for classifying wild lily species, which poses an issue with further characterizing its medicinal properties.    Results  To develop a simple and reliable identification system for  Lilium , 45 representative species from 6 sections were used to develop a DNA barcoding system, which was based on DNA sequence polymorphisms. In this study, we assessed five commonly used DNA barcode candidates ( ITS ,  rbc  L ,  ycf1b ,  mat  K  and  psbA-trnH ) and five novel barcode candidates obtained from highly variable chloroplast genomic regions ( trnL-trnF ,  trnS-trnG ,  trnF-ndhJ ,  trnP-psaJ-rpI33  and  psbB-psbH ). We showed that a set of three novel DNA barcodes ( ITS + trnP-psaJ-rpI33 + psbB-psbH ) could be efficiently used as a genetic marker to distinguish between lily species, as assessed by methods including DNAsp, BI and ML tree, and Pair Wise Group (PWG).    Conclusions  A rapid and reliable DNA barcoding method was developed for all 45 wild  Lilium  species by using  ITS ,  trnP-psaJ-rpI33 , and  psbB-psbH  as DNA barcoding markers. The method can be used in the classification of wild  Lilium  species, especially endangered species, and also provides an effective method for selective lily breeding.    Supplementary Information  The online version contains supplementary material available at 10.1186/s12870-021-03229-6.

Developing an efficient DNA barcoding system to differentiate between Lilium species