FISH Karyotype Analysis Of Wild And Cultural Seabuckthorn Based On 3 Oligo-probes

Seabuckthorn（Hippophae rhamnoides Linn.）,a dioecious deciduous shrub or tree species,has economic and ecological value due to its application in desertification control and medicinal or food.In recent years,oligo probes have been used to label plant chromosomes in fluorescence in situ hybridization（FISH）because of their high cost performance,stable signal and ability to simplify experimental operation.However,FISH based on oligo probes has not been applied to the chromosome labeling of seabuckthorn to explore the differences between wild and cultivated seabuckthorn chromosome structures.At present,there is also a lack of cheap and easy method for gender identification of seabuckthorn.This study for the first time,applied three oligo probes including 5S r DNA,telomere,（TTG）₆ in FISH of 1 kind of wild H.rhamnoides subsp.sinensis material,1 kind of cultural H.rhamnoides subsp.sinensis material,3 kinds of seabuckthorn varieties and an Elaeagnus species.By karyotype analysis and comparing chromosomal morphology differences of various materials,the results were as follows:（1）the telomere signals showed nonspecific signals at both ends of all chromosomes of seabuckthorn and Elaeagnus species.1 pair of interstitial telomere repeat（ITR）signals were located at the primary constrictions of 1 pair of sm chromosomes and 1 pair of m chromosomes of wild seabuckthorn.three pairs（1 strong and 2 weak）（TTG）₆signals were detected in 4 cultural materials,and only 1 pair of relative strong signals were detected in wild H.rhamnoides subsp.sinensis.These signals were distributed at the main constriction,and the strong signals were distributed in 1 pair of SM type chromosomes in cultural materials.（2）Using（TTG）₆ and ITR signals combined with karyotype analysis enabled identifying X chromosomes of the cultural seabuckthorn materials,thus allowing sex identification;the current study discovered heterochromatin structure at chromosome ends in seabuckthorn for the first time,which could be detected by telomere probe signals;Mitotic metaphase chromosomes of endopolyploid cells were detected by telomere signal for the first time,and the frequency of endopolyploid cells differed in different materials;signals of the three probe and heterochromatin allowed specifically identifying of 5chromosomes of wild H.rhamnoides subsp.sinensis;11 chromosomes of cultural H.rhamnoides subsp.sinensis and 10 chromosomes of three kinds of seabuckthorn cultivars respectively.（3）karyotype formulas differed among different materials,but could be explained through FISH karyotyping.Range of chromosome length and total haploid length of chromosome set of wild H.rhamnoides subsp.Sinensis were significantly shorter than those of other cultural materials.（4）The karyotype asymmetry scatter plot shows that the interchromosomal asymmetry of seabuckthorn is low,while the intrachromosomal asymmetry of seabuckthorn is high.FISH based on oligo probes can be applied to identify the difference in chromosome structure and sex of seabuckthorn efficiently,which providing important cytogenetic information for genome assembly and breeding of seabuckthorn.