Comparative analysis of 343 plastid genomes of Solanum section Petota: Insights into potato diversity,phylogeny, and species discrimination

Common potato (Solanum tuberosum L.) and its wild relatives belong to Solanum section Petota. This section's phylogeny and species delimitation are complicated due to various ploidy levels, high heterozygosity, and frequent interspecific hybridization. Compared to the nuclear genome, the plastid genome is more conserved, has a haploid nature, and has a lower nucleotide substitution rate, providing informative alternative insights into the phylogenetic study of section Petota. Here, we analyzed 343 potato plastid genomes from 53 wild and four cultivated species. The diversity of sequences and genomes was comprehensively analyzed. A total of 24 species were placed in a phylogenetic tree based on genomic data for the first time. Overall, our results not only confirmed most existing clades and species boundaries inferred by nuclear evidence but also provided some distinctive species clade belonging and the maternally inherited evidence supporting the hybrid origin of some species. Furthermore, the divergence times between the major potato clades were estimated. In addition, the species discriminatory power of universal barcodes, nuclear ribosomal DNA, and whole and partial plastid genomes and their combinations were thoroughly evaluated; the plastid genome performed best but had limited discriminatory power for all survey species (40%). Overall, our study provided not only new insights into phylogeny and DNA barcoding of potato but also provided valuable genetic data resources for further systematical research of Petota.     

Genomic analyses reveal natural selection on reproduction related genes between two closely related Populus (Salicaceae) species

Identifying the factors that cause reproductive isolation and their relative importance in species divergence is crucial to our understanding of speciation processes. In most species, natural selection is commonly considered to play a large role in driving speciation. Based on whole genome re-sequencing data from 27 Populus alba and 28 Populus adenopoda individuals, we explored the factors related to reproductive isolation of these two closely related species. The results showed that the two species diverged ~5–10 million years ago (Ma), when the Qinghai–Tibet Plateau reached a certain height and the inland climate of the Asian continent became arid. In highly differentiated genomic regions, the relative divergence (F_ST) and absolute divergence (d_xy) were significantly higher than the genomic background, θπ and shared polymorphisms decreased whereas fixed differences increased, which indicated that natural selection played a key role in the reproductive isolation of the two species. In addition, we found several genes that were related to reproduction that may be involved in explaining the reproductive isolation. Using phylogenetic trees resolved from haplotype data of Populus tomentosa and P. adenopoda, the maternal origin of P. tomentosa from P. adenopoda was likely to be located in Hubei and Chongqing Provinces.     

Phylogenomic study of Amorphophallus (Alismatales; Araceae): When plastid DNA gene sequences help to resolve the backbone subgeneric delineation

Encompassing ca. 200 species distributed in paleotropical Africa and Asia, Amorphophallus is one of the largest genera of Araceae. In spite of the great economic interest in its glucomannan production, only a few studies have attempted to grasp the evolutionary history of this genus. In the current state of knowledge, four main clades, mostly linked to biogeographical delineation, have been identified from phylogenies based on a few genes. However, relationships among and within these clades still remain unclear, due to the rapid radiation that occurred during the early evolutionary history of the genus. Here, we generated genome skimming libraries for 43 specimens from 36 species distributed across the 4 clades, which allowed us to produce a phylogenetic matrix for a set of 71 plastid genes. Our phylogenies confirm the monophyly of these clades but show a new and well-resolved arrangement among these clades. Our analyses therefore provide a new scenario and timeline for the evolution of the main Amorphophallus clades, consistent with the morphological characteristics of the clades. The inferred scenario is also in agreement with climate dynamics and the onset of long-distance dispersal by the earliest migratory birds near the Oligocene/Miocene transition around 23 million years ago. Our study provides an up-to-date baseline to understand biogeographic and ecological processes that shaped the current diversity and distribution of Amorphophallus, paving the way for larger-scale phylogenomic studies based on plastid and nuclear genomes.     

EVOLUTIONARY RELATIONSHIPS BETWEEN FOUR SPECIES OF CLADOPHORA (CLADOPHORALES,CHLOROPHYTA) BASED ON DNA–DNA HYBRIDIZATION1

Analysis of the reassociation kinetics of the DNA from Cladophora pellucida (Huds.) Kütz. indicates that the genome of this benthic alga is comprised of approximately 75% repetitive sequences. Single-copy sequences reassociated with a rate constant of 1.8 × 10^?3 M^?1· s^?1, which corresponds to a haploid genome size of 4.7 × 10⁸ bp. Genotypic relationships between members of the form section Longiarticulatae were determined by the method of DNA–DNA hybridization. No significant divergence was observed between the single-copy sequences of C. pellucida isolates from the East Atlantic coast and Mediterranean Sea. Cladophora feredayi Harv. and C. att. ad pellucida from Australia and C. pellucidoidea van den Hoek from the West Atlantic coast were highly and about equally divergent from C. pellucida. The data support the hypothesis that the West Atlantic–West Pacific divergence reflects the middle Miocene closure of the Mediterranean–Indo-Pacific seaways, and the hypothesis that the Northwest Atlantic–Northeast Atlantic divergence reflects the middle Miocene thermal separation of these coasts.     

Construction and characterization of two rice bacterial artificial chromosome libraries from the parents of a permanent recombinant inbred mapping population

Rice is a leading grain crop and the staple food for over half of the world population. Rice is also an ideal species for genetic and biological studies of cereal crops and other monocotyledonous plants because of its small genome and well developed genetic system. To facilitate rice genome analysis leading to physical mapping, the identification of molecular markers closely linked to economic traits, and map-based cloning, we have constructed two rice bacterial artificial chromosome (BAC) libraries from the parents of a permanent mapping population (Lemont and Teqing) consisting of 400 F9 recombinant inbred lines (RILs). Lemont (japonica) and Teqing (indica) represent the two major genomes of cultivated rice, both are leading commercial varieties and widely used germplasm in rice breeding programs. The Lemont library contains 7296 clones with an average insert size of 150 kb, which represents 2.6 rice haploid genome equivalents. The Teqing library contains 14208 clones with an average insert size of 130 kb, which represents 4.4. rice haploid genome equivalents. Three single-copy DNA probes were used to screen the libraries and at least two overlapping BAC clones were isolated with each probe from each library, ranging from 45 to 260 kb in insert size. Hybridization of BAC clones with chloroplast DNA probes and fluorescent in situ hybridization using BAC DNA as probes demonstrated that both libraries contain very few clones of chloroplast DNA origin and are likely free of chimeric clones. These data indicate that both BAC libraries should be suitable for map-based cloning of rice genes and physical mapping of the rice genome.     

Poplar (Populus nigra L.) plants transformed with aBacillus thuringiensis toxin gene: insecticidal activity and genomic analysis

Insect-resistant poplar (Populus nigra L.) plants have been produced by infecting leaves withAgrobacterium tumefaciens strains carrying a binary vector containing different truncated forms of aBacillus thuringiensis (B.t.) toxin gene under a duplicated CaMV 35S promoter. Putative transgenic plants were propagated by cuttings at two experimental farms (in Beijing and Xinjiang, China). At 2–3 years after transformation, 17 of them were selected on the bases of insect-tolerance and good silvicultural traits, and evaluated for insect resistance, for the presence of theB.t. toxin DNA fragment (Southern blots and PCR) and for the expression of the transgene (western and northern blots). Somaclonal variation, as suggested by the appearance of permanent changes in the shape of the leaves, was also investigated with molecular tools (RFLP (restriction fragment length polymorphism), RAPD (random amplified polymorphic DNA) and microsatellite DNA).Bioassays withApochemia cineraius andLymantria dispar on the leaves of the selected clones showed different and, in some cases, high levels of insecticidal activity. The molecular analysis demonstrated integration and expression of the foreign gene. Somatic changes were correlated to extensive genomic changes and were quantified in dendrograms, in terms of genomic similarity. The analysis of control plants suggested that genomic changes were correlated to thein vitro culture step necessary forA. tumefaciens-mediated gene transfer, rather than to the integration of the foreign genes.Three transgenic clones (12, 153 and 192), selected for insect resistance, reduced morphological changes and promising silvicultural traits, are now under large-scale field evaluation in six different provinces in China.     

Cloning and Sequencing of a 27.8-kb Nucleotide Sequence of the 79{degrees}-81{degrees}. Region of the Bacillus subtilis Genome Containing the sspE Locus

The nucleotide sequence of a 27830-bp DNA segment in the 79°–81°.region of the Bacillus subtilis genome has been determined.This region contains 29 complete ORFs including the sspE gene,which encodes a small acid-soluble spore protein gamma and locateson the one side terminal of our assigned region. A homologysearch for the products deduced from the 29 ORFs revealed thatnine of them exhibit significant similarity to known proteins,e.g. proteins involved in an iron uptake system, a multidrugresistance protein, a chloramphenicol resistance protein, epoxidehydrolase, adenine glycosylase, and a glucose-1-dehydrogenasehomolog.     

Rare symmetric and asymmetric Nicotiana tabacum (+) N. megalosiphon somatic hybrids recovered by selection for nuclear-encoded resistance genes and in the absence of genome inactivation

Following protoplast fusion between Nicotiana tabacum (dhfr) and N. megalosiphon (nptII) somatic hybrids were selected on the basis of dual resistance to kanamycin and methotrexate. Despite strong selection for parental nuclear-encoded resistances, only nine N. tabacum (+) N. megalosiphon somatic hybrids were obtained. A preferential loss of the parental N. tabacum nuclear and organelle genome was apparent in some plants in spite of the lack of genomic inactivation by the irradiation or chemical treatment of the parental protoplasts. Only six of the nine hybrids recovered possessed both parental profiles of nuclear RFLPs and isoenzymes. The remaining three hybrids were highly asymmetric with two being identical to N. megalosiphon except for minor morphological differences and rearranged or recombined mitochondrial DNAs (mtDNA), while the other one was distinguishable only by the presence of a rearranged or recombined mtDNA, and was therefore possibly a cybrid. Overall, eight somatic hybrids possessed rearranged or recombined mtDNAs and chloroplast inheritance was non-random since eight possessed N. megalosiphon-type chloroplasts and only one had N. tabacum chloroplasts. In contrast, using the same selection approach, numerous morphologically similar symmetric somatic hybrids with nuclear RFLPs and isozymes of both the parental species were recovered from control fusions between N. tabacum and the more closely related N. sylvestris. In spite of the low frequency of recovery of symmetric N. tabacum (+) N. megalosiphon hybrids in this study, one of these hybrids displayed a significant degree of self-fertility allowing for back-crosses to transfer N. megalosiphon disease-resistance traits to N. tabacum. Plant Research Centre Contribution No. 1579