Genetic Diversity Evaluation And Fingerprint Construction Of Hemerocallis Spp. Based On EST-SSR Molecular Markers

Hemerocallis spp.is distributed all over the world because of its strong adaptability to the environment.The ornamental varieties of daylily are widely used in courtyard and garden greening.With the strengthening of breeding at home and abroad,an increasing number of new horticultural varieties have been cultivated,which makes the genetic structure of daylily more complicated.In this study,new EST-SSR markers were developed by using the transcriptome information of daylily ornamental variety Hemerocallis ‘Autumn Red’ in order to clarify the genetic relationship and promote the sustainable development of breeding of Hemerocallis spp.Then the validity and polymorphism were verified in 104 Hemerocallis accessions including the wild species,varieties and commercial varieties of daylily and landraces of night lily.Finally,30 EST-SSR markers were selected for genetic diversity analysis and DNA fingerprinting construct.The main results of this study were as follows:1.In this study,a total of 28,116 unigenes were detected in the RNA-Seq information of daylily cultivar H.‘Autumn Red’,of which 9048 unigenes contained 9139 SSR loci.One SSR locus was identified every 5.29 kb on average.Tri-nucleotide repeats were predominated(5,270,57.66%)among the SSR loci,followed by di-nucleotide repeats(3,253,35.50%).The number of SSR repeats ranged from 5 to 35,and mainly from 5 to 7,accounting for 70.42%.2.A total of 90 EST-SSR markers were randomly selected to conduct PCR amplification and agarose gel electrophoresis on part of Hemerocallis accessions.Forty valid EST-SSR markers were identified through two rounds of screening,including three pairs of tetranucleotide repeats with five repeat units and 37 pairs of trinucleotide repeats with five to14 repeat units.3.A total of 104 Hemerocallis accessions were amplified and detected by capillary electrophoresis using 40 fluorescent EST-SSR markers.Ninety-Eight alleles were identified from 30 polymorphic markers,with an average value of 3.23.The PIC values ranged from 0.01 to 0.70,with an average of 0.25.Five markers(XC007,XC057,XC058,XC068 and XC072)showed high polymorphism(PIC > 0.5).4.AMOVA analysis of 104 Hemerocallis accessions showed that much of variability resided within accessions.Cluster,STRUCTURE and PCo A analysis showed that 104 accessions were divided into daylily group and night lily group and found that there was gene flow between them.In addition,morphological characteristics of the accessions were combined with molecular markers for analysis.The results showed that the color of flower petals was not enough for classification of Hemerocallis,and there was a significant positive correlation between flower height and flower diameter.We also found that reblooming trait in daylily has potential heritability.These conclusions can provide reference for breeding workers of Hemerocallis.5.DNA fingerprints of 104 Hemerocallis accessions were constructed using seven ESTSSR markers in this study.The Na and PIC values of the seven core markers ranged from 3 to8 and 0.07 to 0.70,respectively.Furthermore,we developed an electronic QR code based on the fingerprint and phenotypic information of 104 germplasm resources.This will help to ensure the authenticity of plant germplasm resources and make easier access to information on varieties by consumers.