Early evolutionary colocalization of the nuclear ribosomal 5S and 45S gene families in seed plants: evidence from the living fossil gymnosperm Ginkgo biloba

In seed plants, the colocalization of the 5S loci within the intergenic spacer (IGS) of the nuclear 45S tandem units is restricted to the phylogenetically derived Asteraceae family. However, fluorescent in situ hybridization (FISH) colocalization of both multigene families has also been observed in other unrelated seed plant lineages. Previous work has identified colocalization of 45S and 5S loci in Ginkgo biloba using FISH, but these observations have not been confirmed recently by sequencing a 1.8 kb IGS. In this work, we report the presence of the 45S–5S linkage in G. biloba, suggesting that in seed plants the molecular events leading to the restructuring of the ribosomal loci are much older than estimated previously. We obtained a 6.0 kb IGS fragment showing structural features of functional sequences, and a single copy of the 5S gene was inserted in the same direction of transcription as the ribosomal RNA genes. We also obtained a 1.8 kb IGS that was a truncate variant of the 6.0 kb IGS lacking the 5S gene. Several lines of evidence strongly suggest that the 1.8 kb variants are pseudogenes that are present exclusively on the satellite chromosomes bearing the 45S–5S genes. The presence of ribosomal IGS pseudogenes best reconciles contradictory results concerning the presence or absence of the 45S–5S linkage in Ginkgo. Our finding that both ribosomal gene families have been unified to a single 45S–5S unit in Ginkgo indicates that an accurate reassessment of the organization of rDNA genes in basal seed plants is necessary

Evolutionary site-number changes of ribosomal DNA loci during speciation: complex scenarios of ancestral and more recent polyploid events

Several genome duplications have been identified in the evolution of seed plants, providing unique systems for studying karyological processes promoting diversification and speciation. Knowledge about the number of ribosomal DNA (rDNA) loci, together with their chromosomal distribution and structure, provides clues about organismal and molecular evolution at various phylogenetic levels. In this work, we aim to elucidate the evolutionary dynamics of karyological and rDNA site-number variation in all known taxa of subtribe Vellinae, showing a complex scenario of ancestral and more recent polyploid events. Specifically, we aim to infer the ancestral chromosome numbers and patterns of chromosome number variation, assess patterns of variation of both 45S and 5S rDNA families, trends in site-number change of rDNA loci within homoploid and polyploid series, and reconstruct the evolutionary history of rDNA site number using a phylogenetic hypothesis as a framework. The best-fitting model of chromosome number evolution with a high likelihood score suggests that the Vellinae core showing x=17 chromosomes arose by duplication events froma recent x=8 ancestor. Our survey suggests more complex patterns of polyploid evolution than previously noted for Vellinae. High polyploidization events (6x, 8x) arose independently in the basal clade Vella castrilensis–V. lucentina, where extant diploid species are unknown. Reconstruction of ancestral rDNA states in Vellinae supports the inference that the ancestral number of loci in the subtribe was two for each multigene family, suggesting that an overall tendency towards a net loss of 5S rDNA loci occurred during the splitting of Vellinae ancestors from the remaining Brassiceae lineages. A contrasting pattern for rDNA site change in both paleopolyploid and neopolyploid species was linked to diversification of Vellinae lineages. This suggests dynamic and independent changes in rDNA site number during speciation processes and a significant lack of correlation between 45S and 5S rDNA evolutionary pathwaysThis research was supported by funds from the Spanish Ministry of Education and Science (Project CGL2010-22347-C02-01), the Catalan Government (Consolidated Research Group 2009SGR608) and by a Ph.D. grant from the Spanish Ministry of Education and Science to J.A.G

Is maize B chromosome preferential fertilization controlled by a single gene?

In previous work, genotypes for high and low B chromosome transmission rate were selected from a native race of maize. It was demonstrated that the B transmission is genetically controlled. The present work reports the fourth and fifth generations of selection and the F1 hybrids between the lines. The native B is characterized by a constant behaviour, with normal meiosis and nondisjunction in 100% of postmeiotic mitosis. It is concluded that genetic variation for B transmission between the selected lines is due to the preferential fertilization process. The F1 hybrids show intermediate B transmission rate between the lines. They are uniform, the variance of the selected character being one order of magnitude lower than that of the native population. In addition, 0B×2B and 2B×2B crosses were made to study the effect of the presence of B chromosomes in the female parent, resulting in non-significant differences. Several crosses were made both in Buenos Aires and in Madrid to compare the possible environmental effect, but significant differences were not found. Our results are consistent with the hypothesis of a single major gene controlling B transmission rate in maize, which acts in the egg cell at the haploid level during fertilization. It is also hypothesized that maize Bs use the normal maize fertilization process to promote their own transmission.Facultad de Ciencias Agrarias y Forestale

Is maize B chromosome preferential fertilization controlled by a single gene?

In previous work, genotypes for high and low B chromosome transmission rate were selected from a native race of maize. It was demonstrated that the B transmission is genetically controlled. The present work reports the fourth and fifth generations of selection and the F1 hybrids between the lines. The native B is characterized by a constant behaviour, with normal meiosis and nondisjunction in 100% of postmeiotic mitosis. It is concluded that genetic variation for B transmission between the selected lines is due to the preferential fertilization process. The F1 hybrids show intermediate B transmission rate between the lines. They are uniform, the variance of the selected character being one order of magnitude lower than that of the native population. In addition, 0B×2B and 2B×2B crosses were made to study the effect of the presence of B chromosomes in the female parent, resulting in non-significant differences. Several crosses were made both in Buenos Aires and in Madrid to compare the possible environmental effect, but significant differences were not found. Our results are consistent with the hypothesis of a single major gene controlling B transmission rate in maize, which acts in the egg cell at the haploid level during fertilization. It is also hypothesized that maize Bs use the normal maize fertilization process to promote their own transmission.Facultad de Ciencias Agrarias y Forestale

Is maize B chromosome preferential fertilization controlled by a single gene?

In previous work, genotypes for high and low B chromosome transmission rate were selected from a native race of maize. It was demonstrated that the B transmission is genetically controlled. The present work reports the fourth and fifth generations of selection and the F1 hybrids between the lines. The native B is characterized by a constant behaviour, with normal meiosis and nondisjunction in 100% of postmeiotic mitosis. It is concluded that genetic variation for B transmission between the selected lines is due to the preferential fertilization process. The F1 hybrids show intermediate B transmission rate between the lines. They are uniform, the variance of the selected character being one order of magnitude lower than that of the native population. In addition, 0B×2B and 2B×2B crosses were made to study the effect of the presence of B chromosomes in the female parent, resulting in non-significant differences. Several crosses were made both in Buenos Aires and in Madrid to compare the possible environmental effect, but significant differences were not found. Our results are consistent with the hypothesis of a single major gene controlling B transmission rate in maize, which acts in the egg cell at the haploid level during fertilization. It is also hypothesized that maize Bs use the normal maize fertilization process to promote their own transmission.Facultad de Ciencias Agrarias y Forestale

Amplification, contraction and genomic spread of a satellite DNA family (E180) in Medicago (Fabaceae) and allied genera

Background and AimsSatellite DNA is a genomic component present in virtually all eukaryotic organisms. The turnover of highly repetitive satellite DNA is an important element in genome organization and evolution in plants. Here we assess the presence and physical distribution of the repetitive DNA E180 family in Medicago and allied genera. Our goals were to gain insight into the karyotype evolution of Medicago using satellite DNA markers, and to evaluate the taxonomic and phylogenetic signal of a satellite DNA family in a genus hypothesized to have a complex evolutionary history.MethodsSeventy accessions from Medicago, Trigonella, Melilotus and Trifolium were analysed by PCR to assess the presence of the repetitive E180 family, and fluorescence in situ hybridization (FISH) was used for physical mapping in somatic chromosomes.Key ResultsThe E180 repeat unit was PCR-amplified in 37 of 40 taxa in Medicago, eight of 12 species of Trigonella, six of seven species of Melilotus and in two of 11 Trifolium species. Examination of the mitotic chromosomes revealed that only 13 Medicago and two Trigonella species showed FISH signals using the E180 probe. Stronger hybridization signals were observed in subtelomeric and interstitial loci than in the pericentromeric loci, suggesting this satellite family has a preferential genomic location. Not all 13 Medicago species that showed FISH localization of the E180 repeat were phylogenetically related. However, nine of these species belong to the phylogenetically derived clade including the M. sativa and M. arborea complexes.ConclusionsThe use of the E180 family as a phylogenetic marker in Medicago should be viewed with caution. Its amplification appears to have been produced through recurrent and independent evolutionary episodes in both annual and perennial Medicago species as well as in basal and derived clades

Genome size, heterochromatin, B chromosome numerical polymorphism and B transmission rate in native races of maize

Se realizaron estudios citogenéticos en poblaciones nativas de maíz del norte argentino adaptadas a diferentes alturas de cultivo. Los mismos consistieron en: determinación del polimorfismo numérico para cromosomas B y del tamaño del genoma, caracterización de la heterocromatina mediante técnicas de bandeos cromosómicos y la relación de éstos con parámetros geográficos como la altitud. Además, se estudió la tasa de transmisión femenina de cromosomas B en una población de la raza Pisingallo. Se estudiaron 21 poblaciones (1120 individuos) cultivadas a diferentes altitudes (80-3620m). Se encontró polimorfismo numérico para cromosomas B en 19 poblaciones. Las frecuencias media de individuos portadores de cromosomas B varían entre 0 y 94%. El análisis de correlación entre el número medio de cromosomas B y la altitud indica que se correlacionan positivamente. El tamaño del genoma (contenido de ADN) fue determinado en 17 poblaciones (107 individuos). Con la finalidad de estudiar la variación del contenido de ADN en los cromosomas A (A-ADN) independientemente de la variación aportada por los Bs, ésta estimación fue llevada a cabo en plantas sin Bs. El rango de variación hallada entre las poblaciones estudiadas fue del 36% (5,00 - 6,8 pg). El contenido de ADN (A-ADN) se correlaciona negativamente con la altura de cultivo y el número medio de Bs. La variación clinal hallada del contenido de ADN y la consecuente correlación con la frecuencia de cromosomas B sobre un gradiente altitudinal tendría un significado adaptativo. En cuatro poblaciones se estudió el contenido de ADN en individuos con diferente dosis de cromosomas B. Se encontró que, dependiendo de la población analizada, la presencia de cromosomas B no siempre incrementa el contenido de ADN respecto a los individuos sin cromosomas B de la misma población. Con el fin de dilucidar sí esto se debe a un efecto de enmascaramiento producido por la variación en el contenido de heterocromatina, se analizaron individuos con distinta dosis de cromosomas B mediante la técnica de bandeo fluorescente utilizando DAPI como fluorocromo. Se encontró, en una población, que la distribución del número de bandas no es independiente de la presencia de cromosomas B. El resultado hallado indica que en individuos portadores de cromosomas B, la variación del número de bandas heterocromáticas estaría enmascarando la presencia de los cromosomas B, al medir el contenido de ADN. Debido a éste resultado existiría una estrecha relación entre el tamaño del genoma, el contenido de heterocromatina y los cromosomas B. En cada población, la frecuencia de Bs podría estar limitada por la variación del contenido de A-ADN o viceversa. Además la distribución de frecuencias en el número de bandas heterocromáticas está relacionada con la presencia de cromosomas B y por lo tanto, los individuos con Bs poseen en general menor número de bandas heterocromáticas en sus cromosomas A, que los individuos sin Bs. Por otro lado, se encontró una correlación altamente significativa entre el número de zonas heterocromáticas y la frecuencia de cromosomas B en 11 poblaciones. Estos resultados sugieren que, en cada población, el contenido de ADN total posee un limite máximo y el mismo sería alcanzado mediante un balance entre el contenido de heterocromatina (en los As) y la dosis de cromosomas B. Por otro lado, con la finalidad de conocer la tasa de transmisión de los cromosomas B por vía materna se realizaron cruzamientos entre plantas f.1B x m. 0B en una población de la raza Pisingallo. Se encontró una variación para la tasa de transmisión de los Bs en la G0. Un experimento de selección con cruzamientos controlados el cual consistió en seleccionar aquellas progenies que presentaron las menores y las mayores tasas de transmisión durante dos generaciones. Los resultados obtenidos en la G1 indican la presencia de dos diferentes grupos de plantas: alta y baja tasa de transmisión de cromosomas B por vía materna y demuestra la existencia de polimorfismo para genes que controlan la tasa de transmisión de los cromosomas B. En la G2 los grupos de alta y baja tasa de transmisión se mantuvieron separados, sin embargo, no fue posible obtener mayor progreso selectivo que el obtenido en la G1. Estos resultados indican que habría un componente genético en la variación obtenida para la tasa de transmisión de los cromosomas B en la generación parental (G0). El grupo seleccionado para alta tasa de transmisión de los cromosomas B por vía materna corresponde a genotipos que lo transmiten de manera mendeliana en cambio, el grupo seleccionado para baja tasa de transmisión lo transmiten a una tasa menor. Es decir que, sólo habría genes "anti-B" que podrían actuar promoviendo la pérdida meiótica del univalente B en la meiósis femenina o la migración del B hacia las megasporas no funcionales o ambos fenómenos. Los mecanismos de acumulación descriptos para maíz son suficiente para explicar el mantenimiento del polimorfismo de los Bs en las poblaciones, sin embargo, estos mecanismos no son suficiente para explicar las diferentes frecuencias de cromosomas B en las poblaciones. Diferentes frecuencias de los alelos que controlan la tasa de transmisión de cromosomas B podrían explicar las frecuencias de cromosomas B hallada en las diferentes poblaciones.Maize native populations from northern Argentina adapted to different altitudes were cytogenetically studied. These studies consisted in: determination of B-chromosome numerical polymorphism, DNA content, characterisation of heterochromatin by techniques of chromosome banding and the study of the relationship with altitude of cultivation. Moreover, the transmission rate of B-chromosome was studied in a population of Pisingallo race. Twenty one native populations (1120 individuals) cultivated at different altitudes (80-3620m) were studied. A numerical polymorphism for B-chromosomes was found in 19 populations. The mean frequencies of individuals with Bs varied from 0 to 94%. A highly significant positive correlation between the mean number of Bs per plant and altitude of cultivation was demonstrated. The genome size (DNA content) was determined in 17 populations (107 individuals). With the aim to study the variation in DNA content of A-chromosomes (A-DNA) independently from the variation supplied by the Bs, this estimation was performed in plants without Bs. The range of variation was 36% (5,00 to 6,8 pg) among studied populations. A-DNA content was negatively correlationed with both altitude of cultivation and mean frequency of Bs. The clinal variation of A-DNA content found in individuals without Bs and the consequent inverse correlation of B frequencies over an altitudinal gradient could have an adaptive significance. The genome size in individuals with different doses of Bs in 4 populations was studied as well. It was found that the presence of Bs did not always increase the DNA content respect to individuals without Bs and it depended on each population. With the aim to know if this phenomenon was caused by a masking effect produced by the variation in heterochromatin amount, individuals with different B-chromosome doses were studied by means of fluorescent banding with DAPI. It was found that the distribution of number of bands is not independent of the presence of B-chromosomes. The found result indicates that in individuals carrying B chromosomes a variation in the number of heterochromatic bands would be masking the B chromosome presence in the DNA content. These results indicate that there is a close interrelationship between the DNA content, heterochromatin amount and B-chromosomes. In each population, the B-chromosome frequency would be limited by the variation of DNA content, or viceversa. Moreover, the distribution of frequency in number of heterochromatic bands is related with the presence of B-chromosomes and, therefore, individuals with Bs have fewer bands than individuals without Bs in the A-chromosomes. On the other hand, a highly significant negative correlation between the number of knobs and the mean number of Bs in eleven populations was found. These results suggest that, in each population, there would be a maximum range of mass of nuclear DNA and this could be reached balancing both heterochromatin amount and B-chromosomes. With the aim to know the transmission rate (TR) of the B chromosomes on the female side, crosses between f.1B X m. 0B plants were carried out in a population (VAV 6313) of the race Pisingallo. Variation of B-chromosome transmission rate in G0 was found. An experiment of selection with controlled crosses was carried out which consisted in selecting those progenies of f.1B X m. 0B crosses showing the highest and lowest TR along two generations. The results obtained in G1 indicate the presence of two different groups of plants, high and low B TR on the female side, and demonstrate the existence of polymorphism for genes controlling B TR. In G2 the high and low B-TR groups were kept up isolated, nevertheless it was not possible to obtain more selective progress than that obtained in G1. This indicates that there is a genetic component to the variation of B TR obtained in G0. The selected groups for high female B-TR corresponds to genotypes that transmit it in a mendelian inheritance, whereas the selected group for low B-TR transmit it at a lower rate tham that in the mendelian inheritance. In other words, there are "anti-B genes" which might act on the propensity for the meiotic loss of the B univalent, or on the nonrandom migration of the B toward the non-functional megaspore, or both. Drive mechanisms are sufficient to account for the maintenance of B polymorphism in maize populations. However, these mechanisms cannot account for the differences of B frequencies in different populations. Different frequencies of the alleles controlling B-TR may account for B frequency in different populations of maize.Fil:Rosato, Carmen Luisa Marcela. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Relationships of the Woody Medicago Species (Section Dendrotelis) Assessed by Molecular Cytogenetic Analyses

†Background and Aims The organization of rDNA genes in the woody medic species from the agronomically important Medicago section Dendrotelis was analysed to gain insight into their taxonomic relationships, to assess the levels of infraspecific variation concerning ribosomal loci in a restricted and fragmented insular species (M. citrina) and to assess the nature of its polyploidy. †Methods Fluorescence in situ hybridization (FISH) was used for physical mapping of 5S and 45S ribosomal DNA genes in the three species of section Dendrotelis (M. arborea, M. citrina, M. strasseri) and the related M. marina from section Medicago. Genomic in situ hybridization (GISH) was used to assess the genomic relationships of the polyploid M. citrina with the putatively related species from section Dendrotelis. †Key Results The diploid (2n 16) M. marina has a single 45S and two 5S rDNA loci, a pattern usually detected in previous studies of Medicago diploid species. However, polyploid species from section Dendrotelis depart from expectations. The tetraploid species (2n 32) M. arborea and M. strasseri have one 45S rDNA locus and two 5S rDNA loci, whereas in the hexaploid (2n 48) M. citrina four 45S rDNA and five 5S rDNA loci have been detected. No single chromosome of M. citrina was uniformly labelled after using genomic probes from M. arborea and M. strasseri. Instead, cross-hybridization signals in M. citrina were restricted to terminal chromo-some arms and NOR regions

Data from: Partial sequence homogenization in the 5S multigene families may generate sequence chimeras and spurious results in phylogenetic reconstructions

Multigene families have provided opportunities for evolutionary biologists to assess molecular evolution processes and phylogenetic reconstructions at deep and shallow systematic levels. However, the use of these markers is not free of technical and analytical challenges. Many evolutionary studies that used the nuclear 5S rDNA gene family rarely used contiguous 5S coding sequences due to the routine use of head-to-tail PCR primers that are anchored to the coding region. Moreover, the 5S coding sequences have been concatenated with independent, adjacent gene units in many studies, creating simulated chimeric genes as the raw data for evolutionary analysis. This practice is based on the tacitly assumed, but rarely tested, hypothesis that strict intra-locus concerted evolution processes are operating in 5S rDNA genes, without any empirical evidence as to whether it holds for the recovered data. The potential pitfalls of analysing the patterns of molecular evolution and reconstructing phylogenies based on these chimeric genes have not been assessed to date. Here, we compared the sequence integrity and phylogenetic behaviour of entire versus concatenated 5S coding regions from a real data set obtained from closely related plant species (Medicago, Fabaceae). Our results suggest that within arrays sequence homogenization is partially operating in the 5S coding region, which is traditionally assumed to be highly conserved. Consequently, concatenating 5S genes increases haplotype diversity, generating novel chimeric genotypes that most likely do not exist within the genome. In addition, the patterns of gene evolution are distorted, leading to incorrect haplotype relationships in some evolutionary reconstructions

Interstitial Telomeric-like Repeats (ITR) in Seed Plants as Assessed by Molecular Cytogenetic Techniques: A Review

The discovery of telomeric repeats in interstitial regions of plant chromosomes (ITRs) through molecular cytogenetic techniques was achieved several decades ago. However, the information is scattered and has not been critically evaluated from an evolutionary perspective. Based on the analysis of currently available data, it is shown that ITRs are widespread in major evolutionary lineages sampled. However, their presence has been detected in only 45.6% of the analysed families, 26.7% of the sampled genera, and in 23.8% of the studied species. The number of ITR sites greatly varies among congeneric species and higher taxonomic units, and range from one to 72 signals. ITR signals mostly occurs as homozygous loci in most species, however, odd numbers of ITR sites reflecting a hemizygous state have been reported in both gymnosperm and angiosperm groups. Overall, the presence of ITRs appears to be poor predictors of phylogenetic and taxonomic relatedness at most hierarchical levels. The presence of ITRs and the number of sites are not significantly associated to the number of chromosomes. The longitudinal distribution of ITR sites along the chromosome arms indicates that more than half of the ITR presences are between proximal and terminal locations (49.5%), followed by proximal (29.0%) and centromeric (21.5%) arm regions. Intraspecific variation concerning ITR site number, chromosomal locations, and the differential presence on homologous chromosome pairs has been reported in unrelated groups, even at the population level. This hypervariability and dynamism may have likely been overlooked in many lineages due to the very low sample sizes often used in cytogenetic studies