| Hawthorn is a kind of Rosaceae plant with the value of ornamental flowers,fruits,and autumn leaves.It is an excellent tree species in the combination of ornamental and production.Hawthorn has a long history of cultivation,variation of species and emergence of varieties are relatively rich.There are 18 species and 6 variations in China.Based on the investigation of germplasm resources in Shenyang and Beijing,The genetic diversity of hawthorn varieties was evaluated from three aspects of morphology,cytology and molecular markers.And the DUS test guidelines for hawthorn new variety,the database of known hawthorn varieties and DNA molecular identification card were developed.The results not only make up for the limitations of traditional research methods,but also provide a scientific basis for the identification,evaluation and utilization of hawthorn variety resources.The main research results are as follows(1)In total,32 phenotypic traits of 108 hawthorn cultivars were studied by means of multivariate statistical analysis.The results showed that the degree of variation was significant,and the genetic diversity was high.Scanning electron microscope was used to observe hawthorn pollen.It was found that hawthorn pollen grains were small,from prolate to perprolate,with three apertures.Pollen shapes and sizes varied among varieties.The types of extine ornamentations were mainly strine-perforated.The chromosome ploidy of hawthorn cultivars was studied by flow cytometry and chromosome tableting techniques.84 diploids,20 triploids and 4 tetraploid were obtained.(2)Using the results of hawthorn transcriptome sequencing obtained by previous research,5091 SSR locus were identified from 14,364 unigenes longer than lkb by MISA software.Among them,dinucleotide repeats are the most,followed by mononucleotides and trinucleotides.The main types of dinucleotide primitives are AG/CT and AAG/CTT,respectively.Primer v3.0 was used to design 500 pairs of SSR primers.After preliminary screening,353(70.60%)primer pairs of the effective amplified products were obtained.After rescreening,304(60.80%)primer pairs with polymorphism were obtained.Finally,24 primer pairs with high polymorphism were selected.(3)24 pairs of specific SSR primers developed from transcriptome were first applied to the study of genetic diversity of hawthorn varieties by multiple fluorescence capillary electrophoresis.A total of 171 alleles were obtained in the study.And the mean value of Na was 7.125.The mean value of Shannon diversity index was 1.178,and the average PIC was 0.550(greater than 0.500),indicating that hawthorn hawthorn had a general level of diversity.(4)The arrangement and combination of 24 pairs of core primers were sorted by serial number,and screening results of primers were encoded in the form of numbers and English letters,forming the DNA molecular ID coding of hawthorn varieties,namely the string DNA molecular identity card.The code is then imported into the online barcode generator to generate barcode DNA molecular identity card.Based on the forms of string and bar code,84 DNA molecular ID of hawthorn varieties were successfully constructed,which is of great significance to the evaluation,utilization and protection of hawthorn varieties resources in China.(5)DUS test guidelines for hawthorn and known variety database in China were developed.There were 55 test characters,including 6 group characters,16 required characters and 12 illustrated characters.And 45 standard varieties were used.Based on the test guidelines,a database of known hawthorn varieties in the form of Excel was constructed,including 108 varieties,each corresponding to 55 phenotypic traits,as well as more than 1,000 photos of the plant,leaves,flowers,fruits and seeds.It provides technical support and scientific basis for the protection of new hawthorn varieties in China. |
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