Preferential elimination of repeated DNA sequences from the paternal, Nicotiana tomentosiformis genome donor of a synthetic, allotetraploid tobacco

Nicotiana tabacum (tobacco, 2n = 4x = 48) is a natural allotetraploid combining two ancestral genomes closely related to modern Nicotiana sylvestris and Nicotiana tomentosiformis. Here we examine the immediate consequences of allopolyploidy on genome evolution using 20 S4-generation plants derived from a single synthetic, S0 plant made by Burk in 1973 (Th37). Using molecular and cytogenetic methods we analysed 14 middle and highly repetitive sequences that together total approximately 4% of the genome. Two repeats related to endogenous geminiviruses (GRD5) and pararetroviruses (NtoEPRV), and two classes of satellite repeats (NTRS, A1/A2) were partially or completely eliminated at variable frequency (25-60%). These sequences are all from the N. tomentosiformis parent. Genomic in situ hybridization revealed additivity in chromosome numbers in two plants (2n = 48), while a third was aneuploid for an N. tomentosiformis-origin chromosome (2n = 49). Two plants had homozygous translocations between chromosomes of the S- and T-genomes. * The data demonstrate that genetic changes in synthetic tobacco were fast, targeted to the paternal N. tomentosiformis-donated genome, and some of the changes showed concordance with changes that presumably occurred during evolution of natural tobacco.     

Long-term genome diploidization in allopolyploid Nicotiana section Repandae (Solanaceae)

Here, we analyze long-term evolution in Nicotiana allopolyploid section Repandae (the closest living diploids are N. sylvestris, the maternal parent, and N. obtusifolia, the paternal parent). We compare data with other more recently formed Nicotiana allopolyploids. We investigated 35S and 5S nuclear ribosomal DNA (rDNA) chromosomal location and unit divergence. A molecular clock was applied to the Nicotiana phylogenetic tree to determine allopolyploid ages. N. tabacum and species of Repandae were c. 0.2 and 4.5 Myr old, respectively. In all Repandae species, the numbers of both 35S and 5S rDNA loci were less than the sum of those of the diploid progenitors. Trees based on 5S rDNA spacer sequences indicated units of only the paternal parent. In recent Nicotiana allopolyploids, the numbers of rDNA loci equal the sum of those of their progenitors. In the Repandae genomes, diploidization is associated with locus loss. Sequence analysis indicates that 35S and 5S units most closely resemble maternal and paternal progenitors, respectively. In Nicotiana, 4.5 Myr of allopolyploid evolution renders genomic in situ hybridization (GISH) unsuitable for the complete resolution of parental genomes.     

First nuclear DNA amounts in more than 300 angiosperms

BACKGROUND AND AIMS: Genome size (DNA C-value) data are key biodiversity characters of fundamental significance used in a wide variety of biological fields. Since 1976, Bennett and colleagues have made scattered published and unpublished genome size data more widely accessible by assembling them into user-friendly compilations. Initially these were published as hard copy lists, but since 1997 they have also been made available electronically (see the Plant DNA C-values database http://www.kew.org/cval/homepage.html). Nevertheless, at the Second Plant Genome Size Meeting in 2003, Bennett noted that as many as 1000 DNA C-value estimates were still unpublished and hence unavailable. Scientists were strongly encouraged to communicate such unpublished data. The present work combines the databasing experience of the Kew-based authors with the unpublished C-values produced by Zonneveld to make a large body of valuable genome size data available to the scientific community. METHODS: C-values for angiosperm species, selected primarily for their horticultural interest, were estimated by flow cytometry using the fluorochrome propidium iodide. The data were compiled into a table whose form is similar to previously published lists of DNA amounts by Bennett and colleagues. KEY RESULTS AND CONCLUSIONS: The present work contains C-values for 411 taxa including first values for 308 species not listed previously by Bennett and colleagues. Based on a recent estimate of the global published output of angiosperm DNA C-value data (i.e. 200 first C-value estimates per annum) the present work equals 1.5 years of average global published output; and constitutes over 12 % of the latest 5-year global target set by the Second Plant Genome Size Workshop (see http://www.kew.org/cval/workshopreport.html). Hopefully, the present example will encourage others to unveil further valuable data which otherwise may lie forever unpublished and unavailable for comparative analyses.     

Blood flows and nutrient uptakes in growth-restricted pregnancies induced by overnourishing adolescent sheep

To establish physiological mechanisms for fetal growth restriction in pregnant adolescent ewes we studied uterine, fetal, and uteroplacental metabolism in ewes offered a high (n = 12) or moderate (n = 10) dietary intake. High intakes decreased placental (226 vs. 414 g, P < 0.001) and fetal weight (3,323 vs. 4,626 g, P < 0.01). Uterine blood flow was reduced absolutely (-36%) but proportional to conceptus weight; umbilical blood flow was reduced absolutely (-37%) and per fetal weight (-15%). Uterine oxygen uptake was decreased per conceptus weight (-14%); there was no change in fetal weight oxygen consumption. Uteroplacental oxygen consumption and clearance were reduced proportional to weight. Similar changes were measured for glucose fluxes and fetal glucose concentration; fetal insulin concentration was reduced. In this model of fetal growth restriction, therefore, maintenance of fetal weight-specific glucose and oxygen consumption rates are producing relative hypoglycemia and hypoxemia. This indicates that increased fetal glucose clearance and/or insulin sensitivity may be operating as compensatory mechanisms to preserve normal fetal metabolism while fetal growth is sacrificed.     

Comparisons with Caenorhabditis (approximately 100 Mb) and Drosophila (approximately 175 Mb) using flow cytometry show genome size in Arabidopsis to be approximately 157 Mb and thus approximately 25% larger than the Arabidopsis genome initiative estimate of approximately 125 Mb

Recent genome sequencing papers have given genome sizes of 180 Mb for Drosophila melanogaster Iso-1 and 125 Mb for Arabidopsis thaliana Columbia. The former agrees with early cytochemical estimates, but numerous cytometric estimates of around 170 Mb imply that a genome size of 125 Mb for arabidopsis is an underestimate. In this study, nuclei of species pairs were compared directly using flow cytometry. Co-run Columbia and Iso-1 female gave a 2C peak for arabidopsis only approx. 15 % below that for drosophila, and 16C endopolyploid Columbia nuclei had approx. 15 % more DNA than 2C chicken nuclei (with >2280 Mb). Caenorhabditis elegans Bristol N2 (genome size approx. 100 Mb) co-run with Columbia or Iso-1 gave a 2C peak for drosophila approx. 75 % above that for 2C C. elegans, and a 2C peak for arabidopsis approx. 57 % above that for C. elegans. This confirms that 1C in drosophila is approx. 175 Mb and, combined with other evidence, leads us to conclude that the genome size of arabidopsis is not approx. 125 Mb, but probably approx. 157 Mb. It is likely that the discrepancy represents extra repeated sequences in unsequenced gaps in heterochromatic regions. Complete sequencing of the arabidopsis genome until no gaps remain at telomeres, nucleolar organizing regions or centromeres is still needed to provide the first precise angiosperm C-value as a benchmark calibration standard for plant genomes, and to ensure that no genes have been missed in arabidopsis, especially in centromeric regions, which are clearly larger than once imagined.     

Evolution of genome size in the angiosperms

Genome size varies extensively across the flowering plants, which has stimulated speculation regarding the ancestral genome size of these plants and trends in genome evolution. We investigated the evolution of C-values across the angiosperms using a molecular phylogenetic framework and C-values not previously available for crucial basal angiosperms, including Amborella, Illiciaceae, and Austrobaileya. Reconstructions of genome size across the angiosperms and extant gymnosperms indicate that the ancestral genome size for angiosperms is very small (1C ≤ 1.4 pg), in agreement with an earlier analysis of Leitch et al. (1998). Furthermore, a very small genome size (1C ≤ 1.4 pg) is ancestral not only for the angiosperms in general, but also for most major clades of flowering plants, including the monocots and the eudicots. The ancestral genome of core eudicots may also have been very small given that very low 1C-values appear to be ancestral for major clades of core eudicots, such as Caryophyllales, Saxifragales, and asterids. Very large genomes occur in clades that occupy derived positions within the monocots and Santalales.     

Rapid evolution of parental rDNA in a synthetic tobacco allotetraploid line

Unidirectional gene conversion of rDNA units has occurred in the evolution of natural tobacco (Nicotiana tabacum). In this paper we report the use of the synthetic tobacco line Th37, 4n (N. sylvestris × N. tomentosiformis), to study early rDNA evolution associated with allopolyploidy. At least three classes of newly amplified rDNA unit variants were identified (17/20 plants). Their presence was often accompanied by near-complete elimination of N. tomentosiformis-donated rDNA units (15/20 plants). Novel rDNA units were of N. tomentosiformis-type and contained rearranged subrepeats in the intergenic spacer. The maternal N. sylvestris-derived units were unchanged, except for some alteration in the ratio of individual gene family members. A cytogenetic analysis revealed rDNA sites on N. sylvestris-derived chromosomes S10, S11, and S12 and N. tomentosiformis-derived chromosomes T3 and in some cases T4. An rDNA locus does not occur on N. tomentosiformis chromosome 4. The locus on chromosome T4 of some hybrids correlates with the occurrence of the novel units that probably amplified at the locus. Combined with an analysis of tobacco cultivars, the data indicate that an initial burst of rDNA evolution associated with allopolyploidy was followed by a slower process that led towards reduced complexity and a decreased number of rDNA variants.     

Evolutionary implications of permanent odd polyploidy in the stable sexual, pentaploid of Rosa canina L

In Rosa canina (2n = 5x = 35), the pollen and ovular parents contribute, respectively, seven and 28 chromosomes to the zygote. At meiosis I, 14 chromosomes form seven bivalents and 21 chromosomes remain as univalents. Fluorescent in situ hybridization to mitotic and pollen mother cells (PMC) of R. canina showed that 10 chromosomes (two per genome) carry ribosomal DNA (rDNA) loci. Five chromosomes carry terminal 18S-5.8S-26S rDNA loci; three of these also carry paracentric 5S rDNA loci and were designated as marker chromosomes 1. Five chromosomes carry only 5S rDNA loci and three of these were designated as marker chromosomes 2. The remaining four of the 10 chromosomes with rDNA loci were individually identifiable by the type and relative sizes of their rDNA loci and were numbered separately. At PMC meiosis, two marker chromosomes 1 and two marker chromosomes 2 formed bivalents, whereas the others were unpaired. In a gynogenetic haploid of R. canina (n = 4x = 28), obtained after pollination with gamma-irradiated pollen, chromosomes at meiosis I in PMC remained predominantly unpaired. The data indicate only one pair of truly homologous genomes in R. canina. The 21 unpaired chromosomes probably remain as univalents through multiple generations and do not recombine. The long-term evolutionary consequence for the univalents is likely to be genetic degradation through accumulated mutational change as in the mammalian Y chromosome and chromosomes of asexual species. But there is no indication that univalents carry degenerate 5S rDNA families. This may point to a recent evolution of the R. canina meiotic system.     

Endogenous gibberellins in immature seeds of Prunus persica L.: identification of GA(118), GA(119), GA(120), GA(121), GA(122) and GA(126)

The endogenous gibberellins in immature seeds of Prunus persica were analyzed by gas chromatography-mass spectrometry. Eleven known gibberellins, GA(3), GA(9), GA(17), GA(19), GA(30), GA(44), GA(61), GA(63), GA(87), GA(95) and GA(97) were identified. Additionally, several hitherto unknown gibberellins were detected and their putative structures were verified by synthesis of the authentic gibberellins. These gibberellins were then assigned trivial numbers, e.g. 1alpha-hydroxy GA(20) (GA(118)), 1alpha-hydroxy GA(9) (GA(119)), 1,2-didehydro GA(9) (GA(120)), 1,2-didehydro GA(70) (GA(121)), 1,2-didehydro GA(69) (GA(122)) and 1,2-didehydro GA(77) (GA(126)). GA(118) and GA(119) were the first 1alpha-hydroxy gibberellins identified from higher plants. The above profile of 1,2-didehydro gibberellins suggests that 1,2-dehydrogenation might occur prior to 3beta-hydroxylation in biosynthesis of GA(3), GA(30) and GA(87) in immature seeds of P. persica.     

Aloe L. – a second plant family without (TTTAGGG)n telomeres

The physical ends of chromosomes are protected and stabilised by telomeres. The sequence of telomeric DNA normally consists of a simple repeating unit that is conserved in many organisms. Most plants examined have been shown to possess Arabidopsis-type telomeres consisting of many repeat copies of the sequence 5′-TTTAGGG-3′. Using fluorescent in situ hybridisation, slot blotting and the asymmetric polymerase chain reaction we demonstrate an absence of Arabidopsis-type telomeres in the genus Aloe (family Asphodelaceae). The only other plant genera so far reported without such telomeres are Allium, Nothoscordum, and Tulbaghia (family Alliaceae). As these genera and Aloe are petaloid monocots in the Asparagales, it is suggested that an absence of Arabidopsis-type telomeres may be characteristic of this related group of plants. Received: 6 September 1999; in revised form: 9 December 1999 / Accepted: 13 December 1999