| Repetitive sequences,including transposable elements(TEs)and tandem repeats(mainly satellite DNAs and microsatellites),occupied a considerable portion of plant genomes.Analysis of the repeat fraction benefits the understanding of genome structure and evolution in plants.Spinach(Spinacia oleracea L.),an important vegetable crop,is also a model dioecious plant species for studying sex determination and sex chromosome evolution.Detailed analysis of its genome repeat sequences can provide important basic data for further study of its genome structure and evolution.In this study,a combination of bioinformatic analysis,fluorescence in situ hybridization(FISH)and other techniques were used to analyze the genome repeatitive sequences and their chromosome localization of spinach.The main results are as follows:1.The transposable elements(TEs)of spinach genome were identified and analyzed by LTRharvest and LTRdigest.A total of 16,002 full-length TEs were identified.The most abundant repeats are long terminal repeats retrotransposons(LTR-REs),and in LTR-REs,Copia elements were overrepresented compared with Gypsy ones.Angela was the most dominating Copia lineage;Ogre/Tat was the most abundant Gypsy lineage.The mean insertion age of LTR-REs was about 1.42 million years;approximately 83.7% of the inserted sequences were retrotransposed during the last two million years.Based on the full-length transposons and other strategies,the TEs of the spinach genome were annotated by RepeatMasker software,and the results showed that transposons accounted for 64.05% of the spinach genome,among which the proportions of LTR-REs,non-LTR-REs,and DNA transposons were 49.2%,2.4% and 5.6%,respectively.FISH analysis showed that most LTR-REs dispersed all over the chromosomes,by contrast,elements of CRM lineage were distributed at the centromeric region of all chromosomes.In addition,Ogre/Tat lineage mainly accumulated on sex chromosomes.2.The original reads of spinach genome was used to identify the satellite DNA by using TAREAN online pipeline.Three satellite DNA sequences were identified: Spsat1,Spsat2 and Spsat3.The lengths of the monomers of the three satellites were 52 bp,365 bp and 325 bp,respectively.FISH analysis indicated that Spsat1 hybridization signal was mainly concentrated in the telomere region of chromosomes 3 and 4,in which the short arm telomere signal of chromosome 3 was relatively strong,while the long arm telomere signal was relatively weak,and the hybridization signal of chromosome 4 was mainly distributed in the long arm telomeres.The hybridization signal of Spsat2 and Spsat3 was located at the end of the long arm of the sex chromosomes.Based on FISH signals of satellites,together with 45 S rDNA,a fine molecular karyotype of spinach was established.Ogre/Tat lineage and the sex chromosome-specific satellites DNAs might participate in sex chromosome formation and evolution.3.In this study,the frequency and distribution of different microsatellites in the spinach genome were characterized.A total of 261,002 perfect microsatellites were identified(estimated frequency: ~262.1 loci/Mbp).The most abundant microsatellites were tetranucleotide and trinucleotide,accounting for 33.2% and 27.7% of the total number of microsatellites,respectively.A total of 105 primer pairs were designed and screened,and 34 were polymorphic among the detected spinach cultivars.Combined with seven primer sets developed previously,41 primer pairs were used to investigate genetic diversity among 43 spinach cultivars in China.The average polymorphism information content value of the 41 markers was 0.43,representing an intermediate level.The spinach cultivars had a low genetic diversity,and no detectable common factors were shared by each group in the UPGMA dendrogram.This study's findings facilitate further investigations on the structure of the spinach genome and provide fundamental data for future breeding applications of spinach in China. |
PDF Full Text Request