Molecular and cytogenetic description of somatic hybrids between Gentiana cruciata L. and G. tibetica King

Karolina Tomiczak

doi:10.1007/s13353-019-00530-x

Abstract

Somatic hybridization provides an opportunity to create cells with new genetic constitution. Here, the interspecific somatic hybrid plants regenerated in vitro following fusion of cell suspension–derived protoplasts of tetraploid Cross Gentian (Gentiana cruciata L., 2n = 52) with protoplasts released from mesophyll tissue of another tetraploid species, Tibetan Gentian (G. tibetica King, 2n = 52), were studied. According to the results of genome analyses with AFLP, ISSR, and CAPS markers, all somatic hybrids were genetically closer to “suspension” fusion partner G. cruciata than to “mesophyll” partner G. tibetica, but they got G. tibetica chloroplasts. Chromosome counting revealed little variation in the number of chromosomes in hybrid’s cells (2n = 88 or 2n = 90), although all plants possessed similar nuclear DNA content which remained stable even after 2 years of in vitro culture. Fluorescence in situ hybridization (FISH) showed that hybrids possessed 4 to 7 chromosomes bearing 5S rDNA sites and 6 or 7 chromosomes with 35S rDNA sites. A part of FISH signals was smaller than those observed in the parental species, which could indicate the loss of rDNA sequences. Genomic in situ hybridization (GISH) showed the predominance of the number of G. cruciata chromosomes over chromosomes of G. tibetica. However, a significant level of cross-hybridization was observed for about one-third of hybrid chromosomes, indicating a high degree of homeology between the genomes of G. cruciata and G. tibetica.

Highlights

Gentiana cruciata L. and Gentiana tibetica King are two endangered tetraploid species of great importance in herbal medicine and horticulture
AFLP analysis of somatic hybrids showed that 191 to 197 amplicons were deleted from G. cruciata, which represented 60.3–62.1% of all bands common with G. cruciata, and 215 to 219 were deleted from
These changes usually occur in a considerably shorter time frame and to a stronger degree than is the case in sexual hybrids (Liu et al 2015). Both AFLP and ISSR banding patterns of somatic hybrids between G. cruciata and G. tibetica showed that about 60% and 70% of bands were deleted from G. cruciata and G. tibetica, respectively

Summary

Introduction

Gentiana cruciata L. and Gentiana tibetica King are two endangered tetraploid species of great importance in herbal medicine and horticulture. Gentiana cruciata (Cross Gentian) is widespread throughout most of Europe and in Western Asia, but it is nowhere very common (Köhlein 1991). Its underground parts (roots and rhizomes) are the source of secoiridoid glycosides such as gentiopicroside, swertiamarine, and sweroside (Szucs et al 2002). These secondary metabolites can be found in roots of plants propagated in vitro (Hayta et al 2011). Gentiana tibetica (Tibetan Gentian) grows in western Nepal, Buthan, and south-eastern Tibet (Köhlein 1991)

Results

Discussion

Conclusion