A. G. Blinov

Molecular phylogeny of the genus Triticum L

The genus Triticum L. includes the major cereal crop, common or bread wheat (hexaploid Triticum aestivum L.), and other important cultivated species. Here, we conducted a phylogenetic analysis of all known wheat species and the closely related Aegilops species. This analysis was based on chloroplast matK gene comparison along with trnL intron sequences of some species. Polyploid wheat species are successfully divided only into two groups – Emmer (sections Dicoccoides and Triticum) and Timopheevii (section Timopheevii). Results reveal strictly maternal plastid inheritance of synthetic wheat amphiploids included in the study. A concordance of chloroplast origin with the definite nuclear genomes of polyploid species that were inherited at the last hybridization events was found. Our analysis suggests that there were two ancestral representatives of Aegilops speltoides Tausch that participated in the speciation of polyploid wheats with B and G genome in their genome composition. However, G genome species are younger in evolution than ones with B genome. B genome-specific PCR primers were developed for amplification of Acc-1 gene.

Population variability in Chironomus (Camptochironomus) species (Diptera, Nematocera) with a Holarctic distribution: evidence of mitochondrial gene flow.

Phylogenetic analysis of DNA sequences from mitochondrial (mt) genes (Cytochrome b and Cytochrome oxidase I) and one nuclear gene (globin 2b) was used for the investigation of Nearctic and Palearctic populations representing four Chironomus species of the subgenus Camptochironomus, namely C. biwaprimus, C. pallidivittatus, C. tentans sensu stricto and C. dilutus (the last two species constitute Holarctic C. tentans sensu lato). Phenograms constructed on the basis of mt sequences were not congruent with trees based on nuclear genes, or with morphological and cytological data. The mt tree divided the populations by continental region, rather than by the species groupings recognized by the other data sets. The incongruence is explained by mt gene flow resulting from hybridization between the sympatric species on each continent. Calculation of divergence times, based on the sequence data, suggest that C. tentans (s.l.) and C. pallidivittatus have both been in North America for about 2.5 My.

Phylogeny of the endemic Baikalian Sergentia (Chironomidae, Diptera)

Fragments of two mitochondrial genes, cytochrome b (CytB) and Cytochrome c oxidase subunit I (COI) have been used as phylogenetic markers in Sergentia (Chironomidae, Diptera). The concatenated (1241 bp) sequences from both genes were used to infer the phylogenetic relationships among seven Sergentia species. Five of the species belong to the endemic fauna of Lake Baikal. Alignments of the nucleotide sequences were used for the construction of trees using Neighbor-Joining and maximum parsimony methods. Both methods yielded similar results. Monophyly of both Sergentia and the Baikalian endemic species was well supported. The date of origin of the endemic group of Sergentia was estimated as 25.7 MYA which closely coincides with the start of geological changes in the Baikal area. A cytological tree, based on 12 chromosomal characteristics, for the same set of Sergentia species showed a great similarity to the molecular phylogeny.

A molecular assessment of the extent of variation and dispersal between Australian populations of the genus Archaeochlus Brundin (Diptera : Chironomidae)

Australian populations of the primitive podonomine genus Archaeochlus have a restricted distribution associated with seepage areas on granite outcrops in south-western and central Australia. These granite outcrops are disjunct and the intervening land offers no apparent suitable habitat for Archaeochlus. This paper uses the DNA sequence of the mitochondrial Cytb gene to assess the phylogenetic relationships between populations of three species, and to examine the hypothesis that these populations achieved their present distribution via ancient river systems that existed up until about 65 million years ago. The phylogeny supports the existence of the three species, but also raises the possibility that a population of A. brundini from Baladjie Rock may be specifically distinct. The phylogenetic relationships do not discriminate between the ancient river hypothesis and alternative explanations. However, estimation of the times of separation of the various populations within species indicates that these populations are too recent to have spread via the river systems. The results show that for each species the most northerly populations are the oldest and that there has been a southward spread of the populations. It is postulated that the southward spread may be a consequence of the increasing aridity of the northern parts of their range since the end of the tertiary.

Molecular phylogeny of theChironomus genus deduced from nucleotide sequences of two nuclear genes,ssp 160 and the globin 2b gene

Two genes were employed to study phylogenetic relatedness of theChironomus species: the protein-coding, salivary gland-specificssp160 gene, and the globin 2b (gb2b) gene. By using PCR, it was demonstrated that all the 38Chironomus species analyzed possess thegb2b gene, while only 13 have thessp160 gene. Partial nucleotide sequences of the genes of 22 species were determined. The data obtained were employed to construct phylogenetic trees which appeared to be topologically similar and revealed five groups of phylogenetically closely related species. Combining the data obtained in the studies of nuclear and mitochondrial genes, a molecular-data-based scenario could be suggested for theChironomus genus evolution.

Insect globin gene polymorphisms: intronic minisatellites and a retroposon interrupting exon 1 of homologous globin genes in Chironomus (Diptera).

Exon 1 of globin gene ct-13RT in clone lambdagb2-1 from Chironomus thummi contains a 444nt SINE (CTRT1). Based on in situ hybridization to polytene salivary gland chromosomes, C. thummi (ct), C. piger (cp) and C. tentans (ctn) contain copies of CTRT1 at multiple chromosomal loci. Genomic PCR amplifications reveal interrupted (ct-13RT) and uninterrupted (ct-13) alleles of the globin gene in the German population of C. thummi maintained in our laboratory, and only uninterrupted alleles or their homologs in different populations of C. thummi, C. piger and C. tentans. PCR amplification did generate different length fragments from cp-13 gene homologs in natural and laboratory C. piger populations that were due to variation in the length of minisatellite expansions of the central introns of the genes rather than a CTRT1-like SINE. Within minisatellite arrays, aligned homologs were more similar than paralogs in a single population, indicating that a tandem cluster of these repeats predated separation of the C. piger populations. The ct-13 genes of several C. thummi populations lack the minisatellites, suggesting their origin in C. piger only after the thummi/piger split. CTRT1 transposition into a ct-13 allele is even more recent, occurring after separation of German and other European C. thummi populations. The nearly intact ct-13RT and comparison with its intact ct-13 allele support a very recent transposition of the CTRT1 SINE into one of at least two already diverged ct-13 globin gene alleles. PCR analysis of DNA from individual adults in C. thummi shows a 1:2:1 distribution of ct-13/ct-13:ct-13/ct-13RT:ct-13RT/ct-13RT genotypes, consistent with a neutral spread of the ct-13RT allele since transposition, and indicating that the hemoglobin encoded by ct-13 is not necessary for survival, at least in a laboratory population of C. thummi.

The Evolution of SINEs and LINEs in the genus Chironomus (Diptera).

Genomic DNA amplification from 51 species of the family Chironomidae shows that most contain relatives of NLRCth1 LINE and CTRT1 SINE retrotransposons first found in Chironomus thummi. More than 300 cloned PCR products were sequenced. The amplified region of the reverse transcriptase gene in the LINEs is intact and highly conserved, suggesting active elements. The SINEs are less conserved, consistent with minimal/no selection after transposition. A mitochondrial gene phylogeny resolves the Chironomus genus into six lineages (Guryev et al. 2001). LINE and SINE phylogenies resolve five of these lineages, indicating their monophyletic origin and vertical inheritance. However, both the LINE and the SINE tree topologies differ from the species phylogeny, resolving the elements into “clusters I–IV” and “cluster V” families. The data suggest a descent of all LINE and SINE subfamilies from two major families. Based on the species phylogeny, a few LINEs and a larger number of SINEs are cladisitically misplaced. Most misbranch with LINEs or SINEs from species with the same families of elements. From sequence comparisons, cladistically misplaced LINEs and several misplaced SINEs arose by convergent base substitutions. More diverged SINEs result from early transposition and some are derived from multiple source SINEs in the same species. SINEs from two species (C. dorsalis, C. pallidivittatus), expected to belong to the clusters I–IV family, branch instead with cluster V family SINEs; apparently both families predate separation of cluster V from clusters I–IV species. Correlation of the distribution of active SINEs and LINEs, as well as similar 3′ sequence motifs in CTRT1 and NLRCth1, suggests coevolving retrotransposon pairs in which CTRT1 transposition depends on enzymes active during NLRCth1 LINE mobility.

Homology-dependent inactivation of LTR retrotransposons in Aspergillus fumigatus and A. nidulans genomes

The repeat-induced point mutation mechanism (RIP) is the most intriguing among the known mechanisms of homology-dependent gene inactivation (silencing) because of its ability to produce irreversible mutations in repetitive DNA sequences. Discovered for the first time in Neurospora crassa, RIP is characterized by C:G to T:A transitions in duplicated sequences. The mechanisms and range of occurrence of RIP are still poorly understood. Mobile elements, including retrotransposons, are a common target for the processes that lead to homology-dependent silencing because of their ability to propagate themselves. Comparative analysis of LTR retrotransposons was performed throughout the genomes of two ascomycetes, Aspergillus fumigatus and A. nidulans. “De-RIP” retroelements were reconstructed on the basis of several copies. CpG, CpA, and TpG sites, which are potential targets for mutagenesis, were found at a much lower frequency in mobile elements than in structural genes. The dinucleotide targets of the two species are affected by RIP at different frequencies: mutagenesis occurs at both CpG and CpA sites in A. fumigatus and is confined to CpG dinucleotides in A. nidulans. This work provides a theoretical background for planning the experimental investigation of RIP inactivation in aspergilli.

Structure and polymorphism of the Chironomus thummi gene encoding special lobe-specific silk protein, ssp160

cDNA encoding Chironomus thummi ssp160 was used to isolate a genomic clone that hybridized in situ to band A2b on polytene chromosome IV, the site of the ssp160 gene. DNA sequencing, primer extension and gene/cDNA nucleotide sequence alignment revealed the gene contains six exons and five introns; 70% of ssp160 is encoded in exon 3. Variations between cDNA and gene sequences led to the design of a polymerase chain reaction, restriction fragment length polymorphism assay that was subsequently used to demonstrate the existence of polymorphic alleles whose distribution varied between geographically separated populations of larvae. The polymorphism is associated with codon deletions in a six-amino-acid repeat containing an N-linked glycosylation motif. These deletions may have resulted from slipped-strand mispairing during DNA replication.

Phylogenetic Relationships among Holarctic Populations of Chironomus entis and Chironomus plumosus in View of Possible Horisontal Transfer of Mitochondrial Genes

In eight Holarctic populations of two typical chironomid sibling species of the plumosus group, Chironomus entisandChironomus plumosus, nucleotide sequences of mitochondrial (cytb) and nuclear (gb2b) gene regions were examined. The phylogenetic trees reflecting the evolutionary histories of the nuclear and mitochondrial markers exhibited significant differences. On the tree based on the nuclear gene sequences the populations clustered according to their species affiliation, whereas on the tree based on the mitochondrial gene sequences the populations were grouped according to their geographic position. This discrepancy is probably explained by mitochondrial gene flow between sympatric species with incomplete reproductive isolation (sibling species). Based on our results together with the earlier data on nuclear and mitochondrial gene sequences of some other species from the phylogenetic group plumosus, a scheme of phylogenetic relationships within this group is proposed. This scheme is in many ways different from the traditional view on the evolutionary relationships among species of the plumosus group.