Chloroplast genome evolution in the genus Avena

The genus Avena contains five different chloroplast genomes, I-V. A physical map of chloroplast (ct) DNA of Avena sativa (type I chloroplast genome) was constructed using three restriction endonucleases, PstI, SalI and SmaI. This genome is ca. 135.5 kbp in size, and contains two inverted repeats of ca. 22.5 kbp each, separated by a large (ca. 79.0 kbp) and small (ca. 12.5 kbp) single copy region. The rbcL gene which codes for the large subunit of ribulose 1,5-bisphosphate carboxylase, was located in the map. Restriction fragment patterns of all five chloroplast genomes were compared, and among them five fragment size and five restriction site mutations were disclosed. Four site mutations were found in two or more chloroplast genomes, the other site and five fragment size mutations were specific to one or another of the chloroplast genomes. A dendrogram showing phylogenetic relationships among the five chloroplast genomes, based on the distribution of the common and specific mutations among them, indicates that chloroplast genome divergence characterized by three restriction site mutations occurred first between two diploid groups, each carrying A and C genome (nuclear), respectively, followed by further speciation in each group.     

Physical map of chloroplast DNA of aerial yam,Dioscorea bulbifera L.

Summary A physical map of chloroplast DNA (ctDNA) of aerial yam, Dioscorea bulbifera L. was constructed using three restriction endonucleases, PstI, SalI, and SmaI. In addition, a clone bank of the BamHI-digested fragments were generated, and the locations of most BamHI fragments on the map were also determined. The ctDNA of D. bulbifera was found to be a circular molecule with a total size of ca. 152 kb involving two inverted repeats of ca. 25.5 kb, and small and large single copy regions of ca. 18.5 and 83.4 kb, respectively. The genes for the large subunit of the ribulose 1,5-bisphosphate carboxylase (rbcL) and the ATP-synthase subunits and (atpB/atpE) were mapped.Contribution from the Plant Germ-plasm Institute and the Laboratory of Genetics (No. 504), Faculty of Agriculture, Kyoto University, Japan. The work was supported in part by a Grant-in-Aid (No. 60400005) from the Ministry of Education, Science and Culture, Japan     

Diversity and evolution of chloroplast DNA in Triticum and Aegilops as revealed by restriction fragment analysis

Summary Restriction fragment analysis of chloroplast (cp) DNAs from 35 wheat (Triticum) and Aegilops species, including their 42 accessions, was carried out with the use of 13 restriction enzymes to clarify variation in their cpDNAs. Fourteen fragment size mutations (deletions/insertions) and 33 recognition site changes were detected among 209 restriction sites sampled. Based on these results, the 42 accessions of wheat-Aegilops could be classified into 16 chloroplast genome types. Most polyploids and their related diploids showed identical restriction fragment patterns, indicating the conservatism of the chloroplast genome during speciation, and maternal lineages of most polyploids were disclosed. This classification of cpDNAs was principally in agreement with that of the plasma types assigned according to phenotypes arising from nucleus-cytoplasm interactions. These mutations detected by restriction fragment analysis were mapped on the physical map of common wheat cpDNA, which was constructed with 13 restriction endonucleases. Length mutations were more frequently observed in some regions than in others: in a 16.0 kilo base pairs (kbp) of DNA region, including rbcL and petA genes, 6 of 14 length mutations were concentrated. This indicates that hot spot regions exist for deletions/insertions in chloroplast genome. On the other hand, 33 recognition site mutations seemed to be distributed equally throughout the genome, except in the inverted repeat region where only one recognition site change was observed. Base substitution rate (p) of cpDNA was similar to that of other plants, such as Brassica, pea and Lycopersicon, showing constant base substitution rates among related taxa and slow evolution of cpDNA compared with animal mitochondrial DNA. Phylogenetic relationships among Triticum and Aegilops species were discussed, based on the present data.Contributions no. 45 and no. 490 from the Kihara Institute for Biological Research, Yokohama City University and the Laboratory of Genetics, Faculty of Agriculture, Kyoto University, respectively.     

The Molecular Basis of Genetic Diversity among Cytoplasms of Triticum and Aegilops. III. Chloroplast Genomes of the M and Modified M Genome-Carrying Species

The restriction fragment patterns of chloroplast DNAs of all M or modified M genome-carrying Aegilops species, and those of common wheat (Triticum aestivum), Ae. umbellulata and Ae. squarrosa as referants, have been analyzed using eight restriction endonucleases, BamHI, EcoRI, HindIII, KpnI, PstI, SalI, SmaI and XhoI. Nine distinctly different chloroplast genomes are evident, and the mutual relatedness among them is estimated based on the number of different restriction fragments. The results lead to the following conclusions. (1) Chloroplast genomes of three Comopyrum species, Ae. comosa, Ae. heldreichii and Ae. uniaristata, are more closely related with each other and are greatly different from those of the Amblyopyrum species, Ae. mutica, and of Ae. umbellulata and Ae. squarrosa. (2) Ae. crassa's chloroplast genome lies at the center of chloroplast genome diversification, whereas those of common wheat, Ae. squarrosa and Ae. uniaristata are three extreme forms lying far from the center. (3) Chloroplast genomes of three 4x species, Ae. biuncialis, Ae. columnaris and Ae. triaristata, arose from Ae. umbellulata, and that of a fourth 4x species, Ae. ventricosa , arose from Ae. squarrosa. The chloroplast origins of two other 4x species, Ae. ovata and Ae. crassa, remain unsolved. (4) The chloroplast genomes of two Ae. mutica strains are identical, even though their cytoplasms exert quite different effects on male fertility, heading date and growth vigor of common wheat.     

RFLP analysis for species separation in the generaBipolaris andCurvularia

Restriction fragment length polymorphism (RFLP) of the total DNA ofBipolaris andCurvularia species was analysed using arbitrarily chosen genomic clones of DNA fromCurvularia lunata andBipolaris maydis as probes. Clear differences among species in both genera, resulting in different banding positions, were obtained with some probe-enzyme combinations. Intraspecific polymorphism in banding positions with these probe-enzyme combinations was slight. These analyses allow discrimination between the species. DNA fingerprinting with intrageneric probes is a potentially useful tool for species separation and identification inBipolaris andCurvularia when coupled with another characteristic such as conidial morphology.Curvularia aeria comb. nov. was proposed forCurvularia lunata var.aeria on the basis of differences in RFLP banding patterns and differences in conidial morphology.     

Whole chloroplast genome comparison of rice,maize, and wheat: implications for chloroplast gene diversification and phylogeny of cereals

The fully sequenced chloroplast genomes of maize (subfamily Panicoideae), rice (subfamily Bambusoideae), and wheat (subfamily Pooideae) provide the unique opportunity to investigate the evolution of chloroplast genes and genomes in the grass family (Poaceae) by whole-genome comparison. Analyses of nucleotide sequence variations in 106 cereal chloroplast genes with tobacco sequences as the outgroup suggested that (1) most of the genic regions of the chloroplast genomes of maize, rice, and wheat have evolved at similar rates; (2) RNA genes have highly conservative evolutionary rates relative to the other genes; (3) photosynthetic genes have been under strong purifying selection; (4) between the three cereals, 14 genes which account for about 28% of the genic region have evolved with heterogeneous nucleotide substitution rates; and (5) rice genes tend to have evolved more slowly than the others at loci where rate heterogeneity exists. Although the mechanism that underlies chloroplast gene diversification is complex, our analyses identified variation in nonsynonymous substitution rates as a genetic force that generates heterogeneity, which is evidence of selection in chloroplast gene diversification at the intrafamilial level. Phylogenetic trees constructed with the variable nucleotide sites of the chloroplast genes place maize basal to the rice-wheat clade, revealing a close relationship between the Bambusoideae and Pooideae.     

Variations in chloroplast proteins and nucleotide sequences of three chloroplast genes in Triticum and Aegilops.

Two alloplasmic wheat lines having the same common wheat nucleus but the cytoplasms of Aegilops crassa and Ae. columnaris together with the corresponding normal line (control) were used in the two-dimensional gel electrophoresis of soluble and thylakoid membrane proteins of the chloroplast. Three chloroplast polypeptides: the Rubisco large subunit, the beta subunit of ATP synthase, and an unidentified 31 kDa protein, differed in the common wheat and two Aegilops cytoplasms. Three chloroplast genes, atpB, atpE and trnM, that respectively encode the beta and epsilon subunits of ATP synthase and tRNA(met), were sequenced. The atpB gene differed by two synonymous base substitutions, whereas the other two genes were identical in the two Aegilops cytoplasms. From the predicted amino acid sequences, the beta subunits of the ATP synthase in the Aegilops cytoplasms were assumed to have three amino acid substitutions: Ala by Val, Asp- by Ala, and Gln by Lys+, in contrast to the cytoplasm of common wheat. This accounts for the difference in pI values found for the common wheat and Aegilops cytoplasms. The two base substitutions for the atpE genes of common wheat and the Aegilops cytoplasms were synonymous. The differences detected in the genes encoding the two subunits of ATP synthase do not appear to be ascribable to the differences in phenotypic effects for the common wheat and Aegilops cytoplasms. The base substitution rate of the atpB-atpE-trnM gene cluster was similar to that of the rbcL gene. From the rate for the atpB gene alone, evolutionary divergence of the wheat-Aegilops complex is assumed to have begun ca. 3.0 x 10(6) years ago, as compared to ca. 8.0 x 10(6) years ago for the divergence of the wheat-Aegilops complex and barley.     

Molecular Variation in Chloroplast DNA Regions in Ancestral Species of Wheat

Restriction map variation in two 5-6-kb chloroplast DNA regions of five diploid Aegilops species in the section Sitopsis and two wild tetraploid wheats, Triticum dicoccoides and Triticum araraticum, was investigated with a battery of four-cutter restriction enzymes. A single accession each of Triticum durum, Triticum timopheevi and Triticum aestivum was included as a reference. More than 250 restriction sites were scored, of which only seven sites were found polymorphic in Aegilops speltoides. No restriction site polymorphisms were detected in all of the other diploid and tetraploid species. In addition, six insertion/deletion polymorphisms were detected, but they were mostly unique or species-specific. Estimated nucleotide diversity was 0.0006 for A. speltoides, and 0.0000 for all the other investigated species. In A. speltoides, none of Tajima's D values was significant, indicating no clear deviation from the neutrality of molecular polymorphisms. Significant non-random association was detected for three combinations out of 10 possible pairs between polymorphic restriction sites in A. speltoides. Phylogenetic relationship among all the plastotypes (plastid genotype) suggested the diphyletic origin of T. dicoccoides and T. araraticum. A plastotype of one A. speltoides accession was identical to the major type of T. araraticum (T. timopheevi inclusively). Three of the plastotypes found in the Sitopsis species are very similar, but not identical, to that of T. dicoccoides, T. durum and T. aestivum.