STAIRS: a new genetic resource for functional genomic studies of Arabidopsis

Many biologically and economically important traits in plants and animals are quantitative/multifactorial, being controlled by several quantitative trait loci (QTL). QTL are difficult to locate accurately by conventional methods using molecular markers in segregating populations, particularly for traits of low heritability or for QTL with small effects. In order to resolve this, large (often unrealistically large) populations are required. In this paper we present an alternative approach using a specially developed resource of lines that facilitate QTL location first to a particular chromosome, then to successively smaller regions within a chromosome (< or = 0.5 cM) by means of simple comparisons among a few lines. This resource consists of "Stepped Aligned Inbred Recombinant Strains" (STAIRS) plus single whole Chromosome Substitution Strains (CSSs). We explain the analytical power of STAIRS and illustrate their construction and use with Arabidopsis thaliana, although the principles could be applied to many organisms. We were able to locate flowering QTL at the top of chromosome 3 known to contain several potential candidate genes.     

A new pheromone trail-based genetic algorithm for comparative genome assembly

Gap closing is considered one of the most challenging and time-consuming tasks in bacterial genome sequencing projects, especially with the emergence of new sequencing technologies, such as pyrosequencing, which may result in large amounts of data without the benefit of large insert libraries for contig scaffolding. We propose a novel algorithm to align contigs with more than one reference genome at a time. This approach can successfully overcome the limitations of low degrees of conserved gene order for the reference and target genomes. A pheromone trail-based genetic algorithm (PGA) was used to search globally for the optimal placement for each contig. Extensive testing on simulated and real data sets shows that PGA significantly outperforms previous methods, especially when assembling genomes that are only moderately related. An extended version of PGA can predict additional candidate connections for each contig and can thus increase the likelihood of identifying the correct arrangement of each contig. The software and test data sets can be accessed at http://sourceforge.net/projects/pga4genomics/.     

Whole genomic DNA sequencing and comparative genomic analysis of Arthrospira platensis: high genome plasticity and genetic diversity

Arthrospira platensis is a multi-cellular and filamentous non-N2-fixing cyanobacterium that is capable of performing oxygenic photosynthesis. In this study, we determined the nearly complete genome sequence of A. platensis YZ. A. platensis YZ genome is a single, circular chromosome of 6.62 Mb in size. Phylogenetic and comparative genomic analyses revealed that A. platensis YZ was more closely related to A. platensis NIES-39 than Arthrospira sp. PCC 8005 and A. platensis C1. Broad gene gains were identified between A. platensis YZ and three other Arthrospira speices, some of which have been previously demonstrated that can be laterally transferred among different species, such as restriction-modification systems-coding genes. Moreover, unprecedented extensive chromosomal rearrangements among different strains were observed. The chromosomal rearrangements, particularly the chromosomal inversions, were analysed and estimated to be closely related to palindromes that involved long inverted repeat sequences and the extensively distributed type IIR restriction enzyme in the Arthrospira genome. In addition, species from genus Arthrospira unanimously contained the highest rate of repetitive sequence compared with the other species of order Oscillatoriales, suggested that sequence duplication significantly contributed to Arthrospira genome phylogeny. These results provided in-depth views into the genomic phylogeny and structural variation of A. platensis, as well as provide a valuable resource for functional genomics studies.     

The first draft genome of Lophophorus: A step forward for Phasianidae genomic diversity and conservation

The monal genus (Lophophorus) is a branch of Phasianidae and its species inhabit the high-altitude mountains of the Qinghai-Tibet Plateau. The Chinese monal, L. lhuysii, is a threatened endemic bird of China that possesses high-altitude adaptability, diversity of plumage color and potentially low reproductive life history. This is the first study to describe the monal genome using next generation sequencing technology. The Chinese monal genome size is 1.01 Gb, with 16,940 protein-coding genes. Gene annotation yielded 100.93 Mb (9.97%) repeat elements, 785 ncRNA, 5,465,549 bp (0.54%) SSR and 15,550 (92%) genes in public databases. Compared to other birds and mammals, the genome evolution analysis showed numerous expanded gene families and positive selected genes involved in high-altitude adaptation, especially related to the adaptation of low temperature and hypoxia. Consequently, this gene data can be used to investigate the molecular evolution of high-altitude adaptation in future bird research. Our first published genome of the genus Lophophorus will be integral for the study of monal population genetic diversity and conservation, genomic evolution and Galliformes species differentiation in the Qinghai-Tibetan Plateau.     

Detailed chromosomal and molecular genetic analysis of single cells by whole genome amplification and comparative genomic hybridisation.

Molecular genetic analysis of isolated single cells and other minute DNA samples is limited because there is insufficient DNA to perform more than one independent PCR amplification. One solution to this problem is to first amplify the entire genome, thus providing enough DNA for numerous subsequent PCRs. In this study we have investigated four different methods of whole genome amplification performed on single cells, and have identified a protocol that generates sufficient quantities of DNA for comparative genomic hybridisation (CGH) as well as more than 90 independent amplification reactions. Thus, numerous specific loci and the copy number of every chromosome can be assessed in a single cell. We report here the first reliable application of CGH to single cells from human preimplantation embryos (blastomeres) and to single fibroblasts, buccal cells and amniocytes.     

Bases and spaces: resources on the web for accessing the draft human genome - II - After publication of the draft

The volume of human genome sequence and the variety of web-based tools to access it continue to grow at an impressive rate, but a working knowledge of certain key resources can be sufficient to get the most from your genome. This article provides an update to Genome Biology 2000, 1(4):reviews2001.1-2001.5.     

Generation of a large-scale genomic resource for functional and comparative genomics in Liriodendron tulipifera L.

Liriodendron tulipifera L., a member of Magnoliaceae in the order Magnoliales, has been used extensively as a reference species in studies on plant evolution. However, genomic resources for this tree species are limited. We constructed cDNA libraries from ten different types of tissues: premeiotic flower buds, postmeiotic flower buds, open flowers, developing fruit, terminal buds, leaves, cambium, xylem, roots, and seedlings. EST sequences were generated either by 454 GS FLX or Sanger methods. Assembly of almost 2.4 million sequencing reads from all libraries resulted in 137,923 unigenes (132,905 contigs and 4,599 singletons). About 50% of the unigenes had significant matches to publically available plant protein sequences, representing a wide variety of putative functions. Approximately 30,000 simple sequence repeats were identified. More than 97% of the cell wall formation genes in the Cell Wall Navigator and the MAIZEWALL databases are represented. The cinnamyl alcohol dehydrogenase (CAD) homologs identified in the L. tulipifera EST dataset showed different expression levels in the ten tissue types included in this study. In particular, the LtuCAD1 was found to partially recover the stiffness of the floral stems in the Arabidopsis thaliana CAD4 and CAD5 double mutant plants, of the LtuCAD1 in lignin biosynthesis. L. tulipifera genes have greater sequence similarity to homologs from other woody angiosperm species than to non-woody model plants. This large-scale genomic resour"HistryDatesce will be instrumental for gene discovery, cDNA microarray production, and marker-assisted breeding in L. tulipifera, and strengthen this species' role in comparative studies.     

Identification of chlorophyllide a oxygenase in the Prochlorococcus genome by a comparative genomic approach

Chl b is a major photosynthetic pigment of peripheral antenna complexes in chlorophytes and prochlorophytes. Chl b is synthesized by chlorophyllide a oxygenase (CAO), an enzyme that has been identified from higher plants, moss, green algae and two groups of prochlorophytes, Prochlorothrix and Prochloron. Based on these results, we previously proposed the hypothesis that all of the Chl b synthesis genes have a common origin. However, the CAO gene is not found in whole genome sequences of Prochlorococcus although a gene which is distantly related to CAO was reported. If Prochlorococcus employs a different enzyme, a Chl synthesis gene should have evolved several times on the different phylogenetic lineages of Prochlorococcus and other Chl b-containing organisms. To examine these hypotheses, we identified a Prochlorococcus Chl b synthesis gene by using a combination of bioinformatics and molecular genetics techniques. We first identified Prochlorococcus-specific genes by comparing the whole genome sequences of Prochlorococcus marinus MED4, MIT9313 and SS120 with Synechococcus sp. WH8102. Synechococcus is closely related to Prochlorococcus phylogenetically, but it does not contain a Chl b synthesis gene. By examining the sequences of Prochlorococcus-specific genes, we found a candidate for the Chl b synthesis gene and introduced it into Synechocystis sp. PCC6803. The transformant cells accumulated Chl b, indicating that the gene product catalyzes Chl b synthesis. In this study, we discuss the evolution of CAO based upon the molecular phylogenetic studies we performed.     

Informatics - genome and genetic databases

Databases for biologists are becoming increasingly important. Some of these can be regarded as ‘core’ resources, such as the bibliographic databases, whereas others are of greater interest to specialists. As comparative genomics develops, however, even databases limited in their scope (e.g. to a single organism) are of great interest to a wider community.     

A comparative genetic map of the turkey genome

Genetic markers (microsatellites and SNPs) were used to create and compare maps of the turkey and chicken genomes. A physical map of the chicken genome was built by comparing sequences of turkey markers with the chicken whole-genome sequence by BLAST analysis. A genetic linkage map of the turkey genome (Meleagris gallopavo) was developed by segregation analysis of genetic markers within the University of Minnesota/Nicholas Turkey Breeding Farms (UMN/NTBF) resource population. This linkage map of the turkey genome includes 314 loci arranged into 29 linkage groups. An additional 40 markers are tentatively placed within linkage groups based on two-point LOD scores and 16 markers remain unlinked. Total map distance contained within linkage groups is 2,011 cM with the longest linkage group (47 loci) measuring 413.3 cM. Average marker interval over the 29 linkage groups was 6.4 cM. All but one turkey linkage group could be aligned with the physical map of the chicken genome. The present genetic map of the turkey provides a comparative framework for future genomic studies.     

Complete genome sequence and comparative genomic analyses of the vancomycin-producing Amycolatopsis orientalis

Background

Amycolatopsis orientalis is the type species of the genus and its industrial strain HCCB10007, derived from ATCC 43491, has been used for large-scale production of the vital antibiotic vancomycin. However, to date, neither the complete genomic sequence of this species nor a systemic characterization of the vancomycin biosynthesis cluster (vcm) has been reported. With only the whole genome sequence of Amycolatopsis mediterranei available, additional complete genomes of other species may facilitate intra-generic comparative analysis of the genus.

Results

The complete genome of A. orientalis HCCB10007 comprises an 8,948,591-bp circular chromosome and a 33,499-bp dissociated plasmid. In total, 8,121 protein-coding sequences were predicted, and the species-specific genomic features of A. orientalis were analyzed in comparison with that of A. mediterranei. The common characteristics of Amycolatopsis genomes were revealed via intra- and inter-generic comparative genomic analyses within the domain of actinomycetes, and led directly to the development of sequence-based Amycolatopsis molecular chemotaxonomic characteristics (MCCs). The chromosomal core/quasi-core and non-core configurations of the A. orientalis and the A. mediterranei genome were analyzed reciprocally, with respect to further understanding both the discriminable criteria and the evolutionary implementation. In addition, 26 gene clusters related to secondary metabolism, including the 64-kb vcm cluster, were identified in the genome. Employing a customized PCR-targeting-based mutagenesis system along with the biochemical identification of vancomycin variants produced by the mutants, we were able to experimentally characterize a halogenase, a methyltransferase and two glycosyltransferases encoded in the vcm cluster. The broad substrate spectra characteristics of these modification enzymes were inferred.

Conclusions

This study not only extended the genetic knowledge of the genus Amycolatopsis and the biochemical knowledge of vcm-related post-assembly tailoring enzymes, but also developed methodology useful for in vivo studies in A. orientalis, which has been widely considered as a barrier in this field.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-363) contains supplementary material, which is available to authorized users.