Rapid genomic DNA changes in allotetraploid fish hybrids

Rapid genomic change has been demonstrated in several allopolyploid plant systems; however, few studies focused on animals. We addressed this issue using an allotetraploid lineage (4nAT) of freshwater fish originally derived from the interspecific hybridization of red crucian carp (Carassius auratus red var., ♀, 2n=100) × common carp (Cyprinus carpio L., ♂, 2n=100). We constructed a bacterial artificial chromosome (BAC) library from allotetraploid hybrids in the 20th generation (F₂₀) and sequenced 14 BAC clones representing a total of 592.126 kb, identified 11 functional genes and estimated the guanine–cytosine content (37.10%) and the proportion of repetitive elements (17.46%). The analysis of intron evolution using nine orthologous genes across a number of selected fish species detected a gain of 39 introns and a loss of 30 introns in the 4nAT lineage. A comparative study based on seven functional genes among 4nAT, diploid F₁ hybrids (2nF₁) (first generation of hybrids) and their original parents revealed that both hybrid types (2nF₁ and 4nAT) not only inherited genomic DNA from their parents, but also demonstrated rapid genomic DNA changes (homoeologous recombination, parental DNA fragments loss and formation of novel genes). However, 4nAT presented more genomic variations compared with their parents than 2nF₁. Interestingly, novel gene fragments were found for the iqca1 gene in both hybrid types. This study provided a preliminary genomic characterization of allotetraploid F₂₀ hybrids and revealed evolutionary and functional genomic significance of allopolyploid animals.     

Genome differentiation in Magonoliaceae as revealed from meiotic pairing in interspecific and intergeneric hybrids

The cross compatibility within and between Yulania Spach and Michelia L.(Magnoliaceae) is relatively good and various such hybrids,obtained by conventional artificial hybridization,are available.The aim of the present study was to determine the extent of genome differentiation between the species involved in these crosses through the observation of chromosome pairing during meiosis in pollen mother cells (PMCs) of the hybrids.Chromosome pairing behavior was studied in five species (2n =38) and two interspecific hybrids of Michelia,eight species (2n =38,76 and 114) and 10 interspecific hybrids of Yulania,and three intergeneric hybrids between Michelia and Yulania.The results showed that chromosome pairing was normal with bivalent formation in diploid parental species and in interspecific hybrids.In addition to bivalents,multivalents were encountered in polyploid parental species and polyploid interspecific hybrids.In the intergeneric hybrids between a tetraploid Yulania and two diploid Michelia,19 chromosomes,most likely originating from Michelia,were unable to synapse from zygotene to metaphase I.Meiotic chromosome pairing indicated a high degree of homology between species within Michelia and Yulania and less homology between the genomes of these two genera.The differentiation of morphological characters and the distinctness of natural distribution also support the conclusion that these two genera are likely independent monophyletic groups.This suggests that the two genera were split at early evolution of Magnoliaceae and the overlapping characteristics in external morphology and internal structures of the two genera may be the result of parallel evolution or ancient common ancestry.     

Production and cytogenetic analysis of intergeneric hybrids betweenElymus anthosachnoides andPsathyrostachys huashanica (Poaceae: Triticeae)

Intergeneric hybridization betweenElymus anthosachnoides (2n = 28,SSYY) andPsathyrostachys huashanica (2n = 14,NN) was performed. Three hybrid plants, obtained via embryo rescue, were intermediate between the parents in morphology and developed vigorously, but were completely sterile. The mean chromosome configuration was 19.48 I + 0.76 II per cell in the hybrids at meiotic metaphase I. This result indicates thatE. anthosachnoides andP. huashanica are distantly related and that there is little or no homoeology betweenN (P. huashanica) andS orY (E. anthosachnoides) genomes.     

DNA fingerprinting of individual species and intergeneric and interspecific hybrids of genera Bos and Bison,subfamily Bovinae

Genome fingerprinting with a hypervariable minisatellite sequence of phage M13 DNA was used to study the genetic variation in individual species of the genera Bos and Bison (subfamily Bovinae) and in their interspecific and intergeneric hybrids. DNA fingerprints were obtained for domestic cow Bos taurus primigenius, vatussy Bos taurus macroceros, banteng Bos javanicus, gaur Bos gaurus, wisent Bison bonasus, bison Bison bison, and for the interspecific and intergeneric hybrids. Compared with the original species, most hybrids showed a greater variation in number and size of hybridization fragments. An association was revealed between the number of hybridization fragments and blood composition of interspecific hybrids resulting from unique crossing of domestic cow and banteng. Pairwise similarity coefficients were calculated to construct a dendrogram of genetic similarity, which reflected the relationships between the parental species and hybrids varying in blood composition. The applicability of the method for identifying interspecific and intergeneric hybrids and for studying the consequences of distant hybridization in the subfamily Bovinae is discussed.     

DNA evidence for multiple origins of intergeneric allopolyploids in annualMicroseris (Asteraceae)

Seventy populations of North American annualMicroseris, Stebbinsoseris, andUropappus species were examined for chloroplast and nuclear ribosomal DNA restriction site variability to determine the origin of the allotetraploid speciesS. heterocarpa andS. decipiens. Previously identified chloroplast DNA restriction site variants were used in concert with restriction site variation forNco I in the nuclear-encoded ribosomal DNA repeat. The presence of two, mutually exclusive restriction site gains were observed in diploid populations ofM. douglasii; these same variants were also found in populations of allotetraploidS. heterocarpa, indicating mutiple origins of this species from different maternal diploid populations ofM. douglasii. Variation in the rDNA repeat between the diploid annual species and the putative paternal genome ofU. lindleyi was found to be additive inS. heterocarpa. A similar relationship was observed for the origin ofS. decipiens; cpDNA restriction site variants found inM. bigelovii andM. douglasii were present inS. decipiens. The rDNANco I variants also were additive in this purported allotetraploid. These results confirm the reticulate evolutionary pattern inStebbinsoseris and provide another example of multiple origins of intergeneric allopolyploids.     

Cytogenetics of new Guizotia Cass. (Compositae) interspecific hybrids pertaining to genomic and phylogenetic affinities

 The following crosses were made using four recognized species/subspecies and a new population of Guizotia, referred to as Chelelu after the name of the locality in Ethiopia from which it was collected: G. scabra subsp. schimperi × Chelelu, Chelelu × G. scabra subsp. scabra, G. zavattarii × G. arborescens and Chelelu × G. zavattarii (all accessions with 2n=30). Plant morphology as well as mitotic and meiotic chromosome analysis confirmed the hybrid nature of the obtained progeny. At metaphase I of meiosis, the F₁ hybrid plants (2n=30) showed a mean of about 95%, 31%, 63% and 0.50% of the pollen mother cells with 15 bivalents, and a mean of about 14.95, 13.75, 14.40 and 7.86 bivalents per cell, respectively. The respective mean pollen stainability was about 67%, 19%, 31% and 2%. From the results it was concluded that Chelelu is more closely related to G. scabra subsp. schimperi than to G. scabra subsp. scabra but more to the latter than to G. zavattarii. Guizotia zavattarii and G. arborescens are closely related to each other. Based on the cytological observations made, the probable basic chromosome number for the genus is discussed. Received November 5, 1999 Accepted August 21, 2001     

Meiotic analysis ofElymus caucasicus,E. longearistatus,and their interspecific hybrids with twenty-threeElymus species (Triticeae,Poaceae)

Interspecific hybridizations were carried out between the two tetraploidsElymus caucasicus andE. longearistatus, and 23 tetraploids and hexaploids ofElymus containing SH, SY, SYH, and SYW genomes and representing various geographical regions. Meiotic pairing was studied in the two target species and their hybrids. It is concluded from this study that (i) interspecific hybridization is fairly easy to perform although strong reproductive barriers exist between the species; (ii)Elymus caucasicus andE. longearistatus are allotetraploids, and share the diverged SY genomes; (iii) the divergence of SY genomes is correlated with the geographic distance between theElymus spp. studied.     

Using genomic slot blot hybridization to assess intergeneric Saccharum x Erianthus hybrids (Andropogoneae - Saccharinae).

The use of genomic slot blot hybridization enabled the differentiation of hybrids from selfs in Saccharum x Erianthus intergeneric crosses in which Saccharum was used as the female parent. Based on the genomic in situ hybridization technique, slot blots of DNA from the parents and the progeny were blocked with the Saccharum parent DNA and hybridized with the labelled male Erianthus genomic DNA. This technique allowed a rapid screening for hybrids and was sensitive enough to detect a 1/20 dilution of Erianthus in Saccharum DNA, which should enable the detection of most partial hybrids. The genomic slot blot hybridization technique was shown to be potentially useful for assessing crosses involving Saccharum species with either Old World Erianthus section Ripidium or North American Erianthus (= Saccharum) species. The effectiveness of the technique was assessed on 144 progeny of a Saccharum officinarum x Erianthus arundinaceus cross, revealing that 43% of the progeny were selfs. The importance of this test as a tool to support intergeneric breeding programs is discussed.     

Genomic instability in sunflower interspecific hybrids

The expression of genomic instability was studied at the phenotypical (morphological characters, electrophoretic spectra of seed storage proteins) and molecular (DNA amplification products) levels in interspecific hybrids (ISHs) from crosses of inbred lines of cultivated sunflower Helianthus annuus with perennial species of the genus Helianthus and in introgressive lines (ILs) produced on their basis. Unstable state of the locus determining the trait of lower branching was proved by the method of hybridological analysis. It was shown with the use of RAPD markers that the IL genome is characterized by instability even after long-term inbreeding (in generations F₈-F₁₂). In progenies of different combinations of interspecific crosses, identical polymorphic variants were revealed for a seed storage protein, helianthinin, and for DNA fragments homologous to structural genes of functionally important proteins, suggesting the nonrandom character of ISH genome variation. This variation may be determined by genome reorganizations under the action of a genome shock induced by interspecific hybridization. The factors inducing reorganizations in the genome include the activity of mobile genetic elements (MGEs). Using primers specific to different MGE families, nucleotide sequences with a high level of homology to the sequences of fragments of the mobile elements MuDR, Far1, CACTA, Stowaway, and Tourist were identified in the sunflower genome. The possibility of using MGE fragments for sunflower genotyping was demonstrated.     

Karyotype stabilization in intergeneric hybrids of the subtribe Triticinae

Summary The C-banded data obtained from Triticinae hybrids are studied with reference to the stabilization of their karyotypes. The types of hybrids distinguished according to genome structure are type I with minimally one diploid genome and type II with a haploid set only. Comparative analysis demonstrates that type I differs from II in karyotype stabilization. The chromosomes from various haploid genomes are combined into new genomes in type I; type II is represented only by amphiploids with the complete set of the chromosomes from all the genomes. The meiotic behaviour of the haploid genome chromosomes were found to have a modifying effect on karyotype stabilization: type II becomes I when homoeologous pairing level is high and when it is associated with the reductional division of univalents.The paper is dedicated to the memory of Professor V. V. Khvostova     

Isolation and characterization of intergeneric somatic hybrids in the Apocynaceae family

Summary Protoplasts were isolated from callus cultures of Rauwolfia serpentina Benth., Rhazya stricta Decaisne, and Catharanthus roseus (L.) G. Don, or from leaves of Vinca minor L. Protoplast isolation, culture, and fusion techniques as well as hybrid screening systems were developed for these species, and hybrids were obtained. Hybrid combinations were Rauwolfia + Vinca, Rauwolfia + Catharanthus, Rauwolfia + Rhazya, and Catharanthus + Vinca. For hybrid isolation, the physiological complementation method was utilized. Analyses of the material obtained included a cytogenetic study of the chromosomes, a study of multiple molecular forms of transferases and esterases, and the blot hybridization of restricted nuclear DNA using ribosomal DNA as a probe. Hybrids were identified in all species' combinations tried. A ten-fold increase in the accumulation of raucaffricine (relative to the parental Rauwolfia strain) was observed in one cell line of the Rauwolfia + Vinca hybrid. Our studies indicated the genetic stability of the great majority of the hybrid cell lines over a period of more than 20 months of in vitro growth. No shoot morphogenesis has so far been observed in this material.