Ty1-copia Reverse Transcriptase Research Articles

Sequence Evolution, Abundance, and Chromosomal Distribution of Ty1-copia Retrotransposons in the Saccharum spontaneum Genome

Saccharum spontaneum is a wild germplasm resource of the genus Saccharum that has many valuable traits. Ty1-copia retrotransposons constitute a large proportion of plant genomes and affect genome sequence organization and evolution. This study aims to analyze the sequence heterogeneity, phylogenetic diversity, copy number, and chromosomal dispersion patterns of Ty1-copia retrotransposons in S. spontaneum. A total of 44 Ty1-copia reverse transcriptase subclones isolated from S. spontaneum showed a range of heterogeneity, and all sequences were A-T rich, averaging approximately 54.59%. Phylogenetic analysis divided the 44 reverse transcriptase sequences into 5 distinct lineages (Retrofit/Ale, Sire/Maximus, Bianca, Tork/TAR, and Ty1-copia like). Dot-blot hybridization revealed that Ty1-copia retrotransposons consisted of a significant component of approximately 38,900 copies and 16,300 copies per genome in the accessions YN82-114 (2n = 10x = 80) and AP85-441 (2n = 4x = 32), respectively. The results of a local blast analysis showed that there are 15,069 Ty1-copia retrotransposon copies in the genome of AP85-441, of which the Retrofit/Ale lineage had the highest copy number, followed by the Tork/TAR, Sire/Maximus, and Bianca lineages. Furthermore, both FISH and the local blast analysis with AP85-441 genomic data demonstrated that the Ty1-copia retrotransposons were unevenly distributed throughout the chromosomes. Taken together, this study provides insights into the role of Ty1-copia retrotransposons in the evolution and organization of the S. spontaneum genome.

Isolation and characterisation of Ty1-copia retrotransposons from Pongamia pinnata

This manuscript mainly deals about the diversity and transcriptional activity of Ty1-copia in Pongamia genome. Pongamia pinnata, a leguminous tree, is popularly known for its biodiesel and medicinal application. However, the structure and the composition of Pongamia nuclear genome are largely unknown. Ty1-copia elements occupy a significant fraction of the plant genome and are responsible for sequence organisation. Here, we have isolated the reverse transcriptase (RT) and RNase H domain of Ty1-copia from Pongamia. In total, 28 Ty1-copia RT sequences were isolated with high levels of sequence heterogeneity. Besides this, the Ty1-copia like elements were also identified in the mitochondrial genome and in the transcriptome library. The analysis revealed that transcriptional activity of Ty1-copia element exists in leaf, root, and seed of salt-stressed and control plants. Dot blot hybridisation reveals that Ty1-copia like retrotransposons occupy around 8.5% of the haploid genome of Pongamia and the phylogenetic study categorised this class of retrotransposons into seven different lineages. These results are useful in understanding the diversity of Ty1-copia lineages and their transcriptional activity present in different tissues of Pongamia.

Characterization, Genomic Organization, Abundance, and Chromosomal Distribution of Ty1-copia Retrotransposons in Erianthus arundinaceus.

Erianthus arundinaceus is an important wild species of the genus Saccharum with many valuable traits. However, the composition and structure of its genome are largely unknown, which have hindered its utilization in sugarcane breeding and evolutionary research. Retrotransposons constitute an appreciable fraction of plant genomes and may have played a significant role in the evolution and sequence organization of genomes. In the current study, we investigate the phylogenetic diversity and genomic abundance of Ty1-copia retrotransposons for the first time and inspect their chromosomal distribution patterns in E. arundinaceus. In total, 70 Ty1-copia reverse transcriptase (RT) sequences with significant levels of heterogeneity were obtained. The phylogenetic analysis revealed these Ty1-copia retrotransposons were classified into four distinct evolutionary lineages (Tork/TAR, Tork/Angela, Retrofit/Ale, and Sire/Maximus). Dot-blot analysis showed estimated the total copy number of Ty1-copia retrotransposons to be about 4.5 × 103 in the E. arundinaceus genome, indicating they were a significant component. Fluorescence in situ hybridization revealed that Ty1-copia retrotransposons from the four lineages had strikingly similar patterns of chromosomal enrichment, being exclusively enriched in the subterminal heterochromatic regions of most E. arundinaceus chromosomes. This is the first clear evidence of the presence of Ty1-copia retrotransposons in the subterminal heterochromatin of E. arundinaceus. Altogether, these results promote the understanding of the diversification of Ty1-copia retrotransposons and shed light on their chromosomal distribution patterns in E. arundinaceus.

LTR-retrotransposon diversity and transcriptional activation under phytoplasma stress in Ziziphus jujuba

Degenerate primers were used to amplify the conserved domains of the Ty1-copia and Ty3-gypsy reverse transcriptase (RT) genes of the long terminal repeat retrotransposons in the jujube (Ziziphus jujuba Mill.) genome. Sequence analysis showed that 18 of the 50 Ty1-copia unique sequences and 28 of the 38 Ty3-gypsy unique sequences possessed stop codons, frameshifts, or both. The overall sequence divergence in jujube was higher in the Ty3-gypsy than the Ty1-copia group, in contrast to the results from other plants. Our results showed that sequence heterogeneity during vertical transmission could be a major influence on the evolution of copia- and gypsy-type retroelements in the jujube genome. In order to elucidate the transcriptional activity under phytoplasma stress of these two classes of RTs, RT-PCR primers were designed to amplify products corresponding to either the Ty3-gypsy or Ty1-copia RT domains from total RNA extracted from leaves or flowers of plants infected by phytoplasma. Amplification products were present for Ty3-gypsy RT in both tissues during infection; however, no RT-PCR products were amplified using the Ty1-copia RT domain primers in phytoplasma-infected leaves or flowers. RT-PCR amplification from total RNA extracted from leaves or flowers of healthy plants did not yield either of the RT fragments. The Ty3-gypsy RT-PCR products were confirmed to originate from the Z. jujuba genome. Our results indicated that the gypsy group of retrotransposons was autonomous retroelements. This is the first report characterizing the Ty1-copia and Ty3-gypsy groups of retrotransposons in the Z. jujuba genome.

Diversity and evolution of Ty1-copia retroelements in representative tribes of Bambusoideae subfamily

Ty1-copia retroelements have been found in all major plants and are largely responsible for the huge differences in the genome size. In this study we isolated and sequenced Ty1-copia reverse transcriptase (rt) gene fragments from 44 representative species of bamboo and nine cultivars or forms of Phyllostachys pubescens. Phylogenetic analysis of 72 distinct Ty1-copia rt sequences showed that Ty1-copia retroelements were widespread, diverse and abundant in these species of Bambusoideae subfamily. In addition, a molecular phylogeny of the species of the Bambusoideae subfamily was established by using the internal transcribed spacer sequences of nuclear ribosomal DNA (ITS) sequences. The comparison between ITS- and Ty1-copia rt- based trees is obviously incongruent. The results suggested either the existence of horizontal transfer events between phylogenetically distant species, or an ancestral Ty1-copia retroelement polymorphism followed by different evolution and stochastic losses.

Phylogenetic and molecular evolutionary analyses of Ty1-copia group retrotransposons in cultivated Egyptian cotton, Gossypium barbadense L.

The Ty1-copia group retrotransposons have been characterized in cultivated Egyptian cotton, G.barbadense L., using degenerate PCR primers for their reverse transcriptase (RT) domains. Comparative nucleotide and amino acid sequences analyses showed that G. barbadense Ty1-copia RT sequences are heterogeneous and this heterogeneity is resolved into 11 distinct families. The high ratio of synonymous to nonsynonymous changes indicates that there is a strong selection for the RT domain of these families. Phylogenetic analysis revealed that two cultivated G. barbadense RT families are closely related to Ty1-copia group retrotransposons present in other plant species. In other words, these families mirror their own phylogenies rather than that of their host cultivars. On the other hand, the remaining G. barbadense RT families are closely related to their respective Gossypium species. These data show that Ty1-copia group retrotransposons tend to span species boundaries, suggesting that they existed early in plant evolution, and were diverged into heterogeneous sequences prior to modern plant species divergence.

Characterisation and physical localisation of Ty1-copia-like retrotransposons in four Alstroemeria species.

The genus Alstroemeria contains species with large genomes (2C = 36.5-78.9 pg (17,600-38,000 Mb) in those species with 2n = 2x = 16). We investigated the diversity and genomic and chromosomal organisation of Ty1-copia-like retrotransposons in four Alstroemeria species. Analysis of 33 PCR-amplified sequences corresponding to a conserved domain of the Ty1-copia reverse transcriptase (rt) gene showed high heterogeneity among predicted amino acid sequences; no two sequences were identical, but most fell into one of five subgroups. Levels of inter- and intra-specific heterogeneity of sequences were similar. HaeIII-digested genomic DNA of various Alstroemeria species contained distinct bands upon hybridisation with individual rt gene fragments. Hybridisation with the heterogeneous PCR pool of rt fragments (retrotransposon pool) revealed additional bands; some minor bands were characteristic of either Brazilian or Chilean species. In situ hybridisation of the retrotransposon pool from three species to metaphase chromosomes from the same species showed a dispersed distribution of the retrotransposon pool with exclusion from rDNA and other chromosomal sites. Alstroemeria pelegrina, which is without major heterochromatic sites, showed some clustering and small negative bands. The retrotransposon pool was excluded from major DAPI-staining bands in Alstroemeria aurea, but in contrast, the sites of the major tandemly repeated sequences in Alstroemeria inodora showed a hybridisation signal similar to that in the rest of the chromosomes. The data are discussed in the context of the contribution of Ty1-copia-like retrotransposons to plant genome size, their evolution, and their value for phylogenetic and biodiversity studies.

Characterisation and physical localisation of Ty1-<i>copia</i>-like retrotransposons in four <i>Alstroemeria</i> species

The genus Alstroemeria contains species with large genomes (2C = 36.5-78.9 pg (17,600-38,000 Mb) in those species with 2n = 2x = 16). We investigated the diversity and genomic and chromosomal organisation of Ty1-copia-like retrotransposons in four Alstroemeria species. Analysis of 33 PCR-amplified sequences corresponding to a conserved domain of the Ty1-copia reverse transcriptase (rt) gene showed high heterogeneity among predicted amino acid sequences; no two sequences were identical, but most fell into one of five subgroups. Levels of inter- and intra-specific heterogeneity of sequences were similar. HaeIII-digested genomic DNA of various Alstroemeria species contained distinct bands upon hybridisation with individual rt gene fragments. Hybridisation with the heterogeneous PCR pool of rt fragments (retrotransposon pool) revealed additional bands; some minor bands were characteristic of either Brazilian or Chilean species. In situ hybridisation of the retrotransposon pool from three species to metaphase chromosomes from the same species showed a dispersed distribution of the retrotransposon pool with exclusion from rDNA and other chromosomal sites. Alstroemeria pelegrina, which is without major heterochromatic sites, showed some clustering and small negative bands. The retrotransposon pool was excluded from major DAPI-staining bands in Alstroemeria aurea, but in contrast, the sites of the major tandemly repeated sequences in Alstroemeria inodora showed a hybridisation signal similar to that in the rest of the chromosomes. The data are discussed in the context of the contribution of Ty1-copia-like retrotransposons to plant genome size, their evolution, and their value for phylogenetic and biodiversity studies.