Identification of stress-responsive genes in an indica rice (Oryza sativa L.) using ESTs generated from drought-stressed seedlings

The impacts of drought on plant growth and development limit cereal crop production worldwide. Rice (Oryza sativa) productivity and production is severely affected due to recurrent droughts in almost all agroecological zones. With the advent of molecular and genomic technologies, emphasis is now placed on understanding the mechanisms of genetic control of the drought-stress response. In order to identify genes associated with water-stress response in rice, ESTs generated from a normalized cDNA library, constructed from drought-stressed leaf tissue of an indica cultivar, Nagina 22 were used. Analysis of 7794 cDNA sequences led to the identification of 5815 rice ESTs. Of these, 334 exhibited no significant sequence homology with any rice ESTs or full-length cDNAs in public databases, indicating that these transcripts are enriched during drought stress. Analysis of these 5815 ESTs led to the identification of 1677 unique sequences. To characterize this drought transcriptome further and to identify candidate genes associated with the drought-stress response, the rice data were compared with those for abiotic stress-induced sequences obtained from expression profiling studies in Arabidopsis, barley, maize, and rice. This comparative analysis identified 589 putative stress-responsive genes (SRGs) that are shared by these diverse plant species. Further, the identified leaf SRGs were compared to expression profiles for a drought-stressed rice panicle library to identify common sequences. Significantly, 125 genes were found to be expressed under drought stress in both tissues. The functional classification of these 125 genes showed that a majority of them are associated with cellular metabolism, signal transduction, and transcriptional regulation

A specific insertion of a solo-LTR characterizes the Y-chromosome of Bryonia dioica (Cucurbitaceae)

Background: Relatively few species of flowering plants are dioecious and even fewer are known to have sex chromosomes. Current theory posits that homomorphic sex chromosomes, such as found in Bryonia dioica (Cucurbitaceae), offer insight into the early stages in the evolution of sex chromosomes from autosomes. Little is known about these early steps, but an accumulation of transposable element sequences has been observed on the Ychromosomes of some species with heteromorphic sex chromosomes. Recombination, by which transposable elements are removed, is suppressed on at least part of the emerging Y-chromosome, and this may explain the correlation between the emergence of sex chromosomes and transposable element enrichment. Findings: We sequenced 2321 bp of the Y-chromosome in Bryonia dioica that flank a male-linked marker, BdY1, reported previously. Within this region, which should be suppressed for recombination, we observed a solo-LTR nested in a Copia-like transposable element. We also found other, presumably paralogous, solo-LTRs in a consensus sequence of the underlying Copia-like transposable element. Conclusions: Given that solo-LTRs arise via recombination events, it is noteworthy that we find one in a genomic region where recombination should be suppressed. Although the solo-LTR could have arisen before recombination was suppressed, creating the male-linked marker BdY1, our previous study on B. dioica suggested that BdY1 may not lie in the recombination-suppressed region of the Y-chromosome in all populations. Presence of a solo-LTR near BdY1 therefore fits with the observed correlation between retrotransposon accumulation and the suppression of recombination early in the evolution of sex chromosomes. These findings further suggest that the homomorphic sex chromosomes of B. dioica, the first organism for which genetic XY sex-determination was inferred, are evolutionarily young and offer reference information for comparative studies of other plant sex chromosomes

Evolution of Genome Size and Complexity in Pinus

BACKGROUND: Genome evolution in the gymnosperm lineage of seed plants has given rise to many of the most complex and largest plant genomes, however the elements involved are poorly understood. METHODOLOGY/PRINCIPAL FINDINGS: Gymny is a previously undescribed retrotransposon family in Pinus that is related to Athila elements in Arabidopsis. Gymny elements are dispersed throughout the modern Pinus genome and occupy a physical space at least the size of the Arabidopsis thaliana genome. In contrast to previously described retroelements in Pinus, the Gymny family was amplified or introduced after the divergence of pine and spruce (Picea). If retrotransposon expansions are responsible for genome size differences within the Pinaceae, as they are in angiosperms, then they have yet to be identified. In contrast, molecular divergence of Gymny retrotransposons together with other families of retrotransposons can account for the large genome complexity of pines along with protein-coding genic DNA, as revealed by massively parallel DNA sequence analysis of Cot fractionated genomic DNA. CONCLUSIONS/SIGNIFICANCE: Most of the enormous genome complexity of pines can be explained by divergence of retrotransposons, however the elements responsible for genome size variation are yet to be identified. Genomic resources for Pinus including those reported here should assist in further defining whether and how the roles of retrotransposons differ in the evolution of angiosperm and gymnosperm genomes

Tomato: a crop species amenable to improvement by cellular and molecular methods

Tomato is a crop plant with a relatively small DNA content per haploid genome and a well developed genetics. Plant regeneration from explants and protoplasts is feasable which led to the development of efficient transformation procedures. In view of the current data, the isolation of useful mutants at the cellular level probably will be of limited value in the genetic improvement of tomato. Protoplast fusion may lead to novel combinations of organelle and nuclear DNA (cybrids), whereas this technique also provides a means of introducing genetic information from alien species into tomato. Important developments have come from molecular approaches. Following the construction of an RFLP map, these RFLP markers can be used in tomato to tag quantitative traits bred in from related species. Both RFLP's and transposons are in the process of being used to clone desired genes for which no gene products are known. Cloned genes can be introduced and potentially improve specific properties of tomato especially those controlled by single genes. Recent results suggest that, in principle, phenotypic mutants can be created for cloned and characterized genes and will prove their value in further improving the cultivated tomato.

The Alcohol Dehydrogenase System in the Xylose-Fermenting Yeast Candida maltosa

The alcohol dehydrogenase (ADH) system plays a critical role in sugar metabolism involving in not only ethanol formation and consumption but also the general "cofactor balance" mechanism. Candida maltosa is able to ferment glucose as well as xylose to produce a significant amount of ethanol. Here we report the ADH system in C. maltosa composed of three microbial group I ADH genes (CmADH1, CmADH2A and CmADH2B), mainly focusing on its metabolic regulation and physiological function.Genetic analysis indicated that CmADH2A and CmADH2B tandemly located on the chromosome could be derived from tandem gene duplication. In vitro characterization of enzymatic properties revealed that all the three CmADHs had broad substrate specificities. Homo- and heterotetramers of CmADH1 and CmADH2A were demonstrated by zymogram analysis, and their expression profiles and physiological functions were different with respect to carbon sources and growth phases. Fermentation studies of ADH2A-deficient mutant showed that CmADH2A was directly related to NAD regeneration during xylose metabolism since CmADH2A deficiency resulted in a significant accumulation of glycerol.Our results revealed that CmADH1 was responsible for ethanol formation during glucose metabolism, whereas CmADH2A was glucose-repressed and functioned to convert the accumulated ethanol to acetaldehyde. To our knowledge, this is the first demonstration of function separation and glucose repression of ADH genes in xylose-fermenting yeasts. On the other hand, CmADH1 and CmADH2A were both involved in ethanol formation with NAD regeneration to maintain NADH/NAD ratio in favor of producing xylitol from xylose. In contrast, CmADH2B was expressed at a much lower level than the other two CmADH genes, and its function is to be further confirmed

Detailed Analysis of a Contiguous 22-Mb Region of the Maize Genome

Most of our understanding of plant genome structure and evolution has come from the careful annotation of small (e.g., 100 kb) sequenced genomic regions or from automated annotation of complete genome sequences. Here, we sequenced and carefully annotated a contiguous 22 Mb region of maize chromosome 4 using an improved pseudomolecule for annotation. The sequence segment was comprehensively ordered, oriented, and confirmed using the maize optical map. Nearly 84% of the sequence is composed of transposable elements (TEs) that are mostly nested within each other, of which most families are low-copy. We identified 544 gene models using multiple levels of evidence, as well as five miRNA genes. Gene fragments, many captured by TEs, are prevalent within this region. Elimination of gene redundancy from a tetraploid maize ancestor that originated a few million years ago is responsible in this region for most disruptions of synteny with sorghum and rice. Consistent with other sub-genomic analyses in maize, small RNA mapping showed that many small RNAs match TEs and that most TEs match small RNAs. These results, performed on ∼1% of the maize genome, demonstrate the feasibility of refining the B73 RefGen_v1 genome assembly by incorporating optical map, high-resolution genetic map, and comparative genomic data sets. Such improvements, along with those of gene and repeat annotation, will serve to promote future functional genomic and phylogenomic research in maize and other grasses

The Selaginella Genome Identifies Genetic Changes Associated with the Evolution of Vascular Plants

Vascular plants appeared ~410 million years ago then diverged into several lineages of which only two survive: the euphyllophytes (ferns and seed plants) and the lycophytes (1). We report here the genome sequence of the lycophyte Selaginella moellendorffii (Selaginella), the first non-seed vascular plant genome reported. By comparing gene content in evolutionary diverse taxa, we found that the transition from a gametophyte- to sporophyte- dominated life cycle required far fewer new genes than the transition from a non-seed vascular to a flowering plant, while secondary metabolic genes expanded extensively and in parallel in the lycophyte and angiosperm lineages. Selaginella differs in post- transcriptional gene regulation, including small RNA regulation of repetitive elements, an absence of the tasiRNA pathway and extensive RNA editing of organellar genes

Control of flowering time and spike development in cereals: the earliness per se Eps-1 region in wheat, rice, and Brachypodium

The earliness per se gene Eps-Am1 from diploid wheat Triticum monococcum affects heading time, spike development, and spikelet number. In this study, the Eps1 orthologous regions from rice, Aegilops tauschii, and Brachypodium distachyon were compared as part of current efforts to clone this gene. A single Brachypodium BAC clone spanned the Eps-Am1 region, but a gap was detected in the A. tauschii physical map. Sequencing of the Brachypodium and A. tauschii BAC clones revealed three genes shared by the three species, which showed higher identity between wheat and Brachypodium than between them and rice. However, most of the structural changes were detected in the wheat lineage. These included an inversion encompassing the wg241-VatpC region and the presence of six unique genes. In contrast, only one unique gene (and one pseudogene) was found in Brachypodium and none in rice. Three genes were present in both Brachypodium and wheat but were absent in rice. Two of these genes, Mot1 and FtsH4, were completely linked to the earliness per se phenotype in the T. monococcum high-density genetic map and are candidates for Eps-Am1. Both genes were expressed in apices and developing spikes, as expected for Eps-Am1 candidates. The predicted MOT1 protein showed amino acid differences between the parental T. monococcum lines, but its effect is difficult to predict. Future steps to clone the Eps-Am1 gene include the generation of mot1 and ftsh4 mutants and the completion of the T. monococcum physical map to test for the presence of additional candidate genes

A BAC pooling strategy combined with PCR-based screenings in a large, highly repetitive genome enables integration of the maize genetic and physical maps

BACKGROUND: Molecular markers serve three important functions in physical map assembly. First, they provide anchor points to genetic maps facilitating functional genomic studies. Second, they reduce the overlap required for BAC contig assembly from 80 to 50 percent. Finally, they validate assemblies based solely on BAC fingerprints. We employed a six-dimensional BAC pooling strategy in combination with a high-throughput PCR-based screening method to anchor the maize genetic and physical maps. RESULTS: A total of 110,592 maize BAC clones (~ 6x haploid genome equivalents) were pooled into six different matrices, each containing 48 pools of BAC DNA. The quality of the BAC DNA pools and their utility for identifying BACs containing target genomic sequences was tested using 254 PCR-based STS markers. Five types of PCR-based STS markers were screened to assess potential uses for the BAC pools. An average of 4.68 BAC clones were identified per marker analyzed. These results were integrated with BAC fingerprint data generated by the Arizona Genomics Institute (AGI) and the Arizona Genomics Computational Laboratory (AGCoL) to assemble the BAC contigs using the FingerPrinted Contigs (FPC) software and contribute to the construction and anchoring of the physical map. A total of 234 markers (92.5%) anchored BAC contigs to their genetic map positions. The results can be viewed on the integrated map of maize [1,2]. CONCLUSION: This BAC pooling strategy is a rapid, cost effective method for genome assembly and anchoring. The requirement for six replicate positive amplifications makes this a robust method for use in large genomes with high amounts of repetitive DNA such as maize. This strategy can be used to physically map duplicate loci, provide order information for loci in a small genetic interval or with no genetic recombination, and loci with conflicting hybridization-based information