John Jacobs

CMD: a Cotton Microsatellite Database resource for Gossypium genomics

BackgroundThe Cotton Microsatellite Database (CMD) http://www.cottonssr.org is a curated and integrated web-based relational database providing centralized access to publicly available cotton microsatellites, an invaluable resource for basic and applied research in cotton breeding.DescriptionAt present CMD contains publication, sequence, primer, mapping and homology data for nine major cotton microsatellite projects, collectively representing 5,484 microsatellites. In addition, CMD displays data for three of the microsatellite projects that have been screened against a panel of core germplasm. The standardized panel consists of 12 diverse genotypes including genetic standards, mapping parents, BAC donors, subgenome representatives, unique breeding lines, exotic introgression sources, and contemporary Upland cottons with significant acreage. A suite of online microsatellite data mining tools are accessible at CMD. These include an SSR server which identifies microsatellites, primers, open reading frames, and GC-content of uploaded sequences; BLAST and FASTA servers providing sequence similarity searches against the existing cotton SSR sequences and primers, a CAP3 server to assemble EST sequences into longer transcripts prior to mining for SSRs, and CMap, a viewer for comparing cotton SSR maps.ConclusionThe collection of publicly available cotton SSR markers in a centralized, readily accessible and curated web-enabled database provides a more efficient utilization of microsatellite resources and will help accelerate basic and applied research in molecular breeding and genetic mapping in Gossypium spp.

Meta-analysis of cotton fiber quality QTLs across diverse environments in a Gossypium hirsutum x G. barbadense RIL population

BackgroundCotton fibers (produced by Gossypium species) are the premier natural fibers for textile production. The two tetraploid species, G. barbadense (Gb) and G. hirsutum (Gh), differ significantly in their fiber properties, the former having much longer, finer and stronger fibers that are highly prized. A better understanding of the genetics and underlying biological causes of these differences will aid further improvement of cotton quality through breeding and biotechnology. We evaluated an inter-specific Gh × Gb recombinant inbred line (RIL) population for fiber characteristics in 11 independent experiments under field and glasshouse conditions. Sites were located on 4 continents and 5 countries and some locations were analyzed over multiple years.ResultsThe RIL population displayed a large variability for all major fiber traits. QTL analyses were performed on a per-site basis by composite interval mapping. Among the 651 putative QTLs (LOD > 2), 167 had a LOD exceeding permutation based thresholds. Coincidence in QTL location across data sets was assessed for the fiber trait categories strength, elongation, length, length uniformity, fineness/maturity, and color. A meta-analysis of more than a thousand putative QTLs was conducted with MetaQTL software to integrate QTL data from the RIL and 3 backcross populations (from the same parents) and to compare them with the literature. Although the global level of congruence across experiments and populations was generally moderate, the QTL clustering was possible for 30 trait x chromosome combinations (5 traits in 19 different chromosomes) where an effective co-localization of unidirectional (similar sign of additivity) QTLs from at least 5 different data sets was observed. Most consistent meta-clusters were identified for fiber color on chromosomes c6, c8 and c25, fineness on c15, and fiber length on c3.ConclusionsMeta-analysis provided a reliable means of integrating phenotypic and genetic mapping data across multiple populations and environments for complex fiber traits. The consistent chromosomal regions contributing to fiber quality traits constitute good candidates for the further dissection of the genetic and genomic factors underlying important fiber characteristics, and for marker-assisted selection.

An active role for endogenous β‐1,3‐glucanase genes in transgene‐mediated co‐suppression in tobacco

Post‐transcriptional gene silencing (PTGS) is characterized by the accumulation of short interfering RNAs that are proposed to mediate sequence‐specific degradation of cognate and secondary target mRNAs. In plants, it is unclear to what extent endogenous genes contribute to this process. Here, we address the role of the endogenous target genes in transgene‐mediated PTGS of β‐1,3‐glucanases in tobacco. We found that mRNA sequences of the endogenous glucanase glb gene with varying degrees of homology to the Nicotiana plumbaginifolia gn1 transgene are targeted by the silencing machinery, although less efficiently than corresponding transgene regions. Importantly, we show that endogene‐specific nucleotides in the glb sequence provide specificity to the silencing process. Consistent with this finding, small sense and antisense 21‐ to 23‐nucleotide RNAs homologous to the endogenous glb gene were detected. Combined, these data demonstrate that a co‐suppressed endogenous glucan ase gene is involved in signal amplification and selection of homologous targets, and show that endogenous genes can actively participate in PTGS in plants. The findings are introduced as a further sophistication of the post‐transciptional silencing model.

Deep Sequencing Reveals Differences in the Transcriptional Landscapes of Fibers from Two Cultivated Species of Cotton

Cotton (Gossypium) fiber is the most prevalent natural product used in the textile industry. The two major cultivated species, G. hirsutum (Gh) and G. barbadense (Gb), are allotetraploids with contrasting fiber quality properties. To better understand the molecular basis for their fiber differences, EST pyrosequencing was used to document the fiber transcriptomes at two key development stages, 10 days post anthesis (dpa), representing the peak of fiber elongation, and 22 dpa, representing the transition to secondary cell wall synthesis. The 617,000 high quality reads (89% of the total 692,000 reads) from 4 libraries were assembled into 46,072 unigenes, comprising 38,297 contigs and 7,775 singletons. Functional annotation of the unigenes together with comparative digital gene expression (DGE) revealed a diverse set of functions and processes that were partly linked to specific fiber stages. Globally, 2,770 contigs (7%) showed differential expression (>2-fold) between 10 and 22 dpa (irrespective of genotype), with 70% more highly expressed at 10 dpa, while 2,248 (6%) were differentially expressed between the genotypes (irrespective of stage). The most significant genes with differential DGE at 10 dpa included expansins and lipid transfer proteins (higher in Gb), while at 22 dpa tubulins, cellulose, and sucrose synthases showed higher expression in Gb. DGE was compared with expression data of 10 dpa-old fibers from Affymetrix microarrays. Among 543 contigs showing differential expression on both platforms, 74% were consistent in being either over-expressed in Gh (242 genes) or in Gb (161 genes). Furthermore, the unigene set served to identify 339 new SSRs and close to 21,000 inter-genotypic SNPs. Subsets of 88 SSRs and 48 SNPs were validated through mapping and added 65 new loci to a RIL genetic map. The new set of fiber ESTs and the gene-based markers complement existing available resources useful in basic and applied research for crop improvement in cotton.

Heritability and predicted selection response of yield components and fibre properties in an inter-specific derived RIL population of cotton

Exploiting genetic variation through inter-specific breeding has improved cotton yield, fibre properties and adaptability. The objectives of this study were to examine heritability and predicted selection response of yield components and fibre properties in a recombinant inbred line (RIL) population from an inter-specific cross between Gossypium hirsutum (Gh) variety Guazuncho 2, and G. barbadense (Gb) line VH8-4602. A population of 93 and 82 RILs was tested in two seasons, with two parents and local controls, Sicot 75 (Gh) and Sipima 280 (Gb) in field experiments. Seed cotton samples hand harvested before and after defoliation were used to measure lint percent, boll weight, 100 seed weight and the lint to measure fibre length, uniformity, short fibre index (SFI), elongation, strength, micronaire, maturity ratio (MR), percent of maturity (PM) and fineness. There was large phenotypic variation for individual traits and transgressive segregation occurred in lint percent, lint weight/seed, fibre no./seed, uniformity, SFI, elongation, MR and PM. Narrow sense heritabilities were moderate for yield components (34.3–41.2%) and for key fibre properties, length, strength, micronaire and fineness (38.3–42.1%), which led to a predicted selection response of 6.7–24.0% for yield components and 3.9–10.9% for key fibre properties under a selection intensity of 10%. Favourable associations were found between key fibre properties, but an adverse association between lint percent and each of these fibre properties. Only five RILs were identified with desirable combinations. The results demonstrated the value of exploiting inter-specific variation to develop cotton germplasm and how breeding strategies can be improved.

Identification of positive and negative regulators of disease resistance to rice blast fungus using constitutive gene expression patterns.

Elevated constitutive expression of components of the defence arsenal is associated with quantitative resistance to the rice blast fungus, a phenomenon called preformed defence. While the role of many disease regulators in inducible defence systems has been extensively studied, little attention has been paid so far to genes that regulate preformed defence. In this study, we show by microarray analysis across rice diversity that the preformed defence phenomenon impacts on a large number of defence-related genes without apparently affecting other biological processes. Using a guilt-by-association strategy, we identified two positive regulators that promote constitutive expression of known defence markers and partial resistance to rice blast. The HSF23 gene encodes for a putative member of the heat shock transcription factor family, while CaMBP encodes for a putative Calmodulin-binding protein. Both HSF23 and CaMBP strongly affect preformed defence and also plant growth. Additionally, we identified the OB-fold gene as a negative regulator of blast resistance, which could be involved in RNA stabilization. The OB-fold mutants do not suffer from obvious developmental defects. Taken together, our results prove that our strategy of combining analysis of gene expression diversity with guilt-by-association is a powerful way to identify disease resistance regulators in rice.