Brent Pedersen

Finding and Comparing Syntenic Regions among Arabidopsis and the Outgroups Papaya, Poplar, and Grape: CoGe with Rosids

In addition to the genomes of Arabidopsis (Arabidopsis thaliana) and poplar (Populus trichocarpa), two near-complete rosid genome sequences, grape (Vitis vinifera) and papaya (Carica papaya), have been recently released. The phylogenetic relationship among these four genomes and the placement of their three independent, fractionated tetraploidies sum to a powerful comparative genomic system. CoGe, a platform of multiple whole or near-complete genome sequences, provides an integrative Web-based system to find and align syntenic chromosomal regions and visualize the output in an intuitive and interactive manner. CoGe has been customized to specifically support comparisons among the rosids. Crucial facts and definitions are presented to clearly describe the sorts of biological questions that might be answered in part using CoGe, including patterns of DNA conservation, accuracy of annotation, transposability of individual genes, subfunctionalization and/or fractionation of syntenic gene sets, and conserved noncoding sequence content. This précis of an online tutorial, CoGe with Rosids (http://tinyurl.com/4a23pk), presents sample results graphically.

Following Tetraploidy in Maize, a Short Deletion Mechanism Removed Genes Preferentially from One of the Two Homeologs

Following genome duplication and selfish DNA expansion, maize used a heretofore unknown mechanism to shed redundant genes and functionless DNA with bias toward one of the parental genomes.

Many or most genes in Arabidopsis transposed after the origin of the order Brassicales.

Previous to this work, typical genes were thought to move from one position to another infrequently. On the contrary, we now estimate that between one-fourth and three-fourths of the genes in Arabidopsis transposed in the Brassicales. We used the CoGe comparative genomics system to perform and visualize multiple orthologous chromosomal alignments. Using this tool, we found large differences between different categories of genes. Ten of the gene families examined, including genes in most transcription factor families, exhibited a median frequency of 5% transposed genes. In contrast, other gene families were composed largely of transposed genes: NB-LRR disease-resistance genes, genes encoding MADS-box and B3 transcription factors, and genes encoding F-box proteins. A unique method involving transposition-rich regions of genome allowed us to obtain an indirect estimate of the positional stability of the average gene. The observed differences between gene families raise important questions concerning the causes and consequences of gene transposition.

The Value of Nonmodel Genomes and an Example Using SynMap Within CoGe to Dissect the Hexaploidy that Predates the Rosids

We find great value in the genomes from the nonmodel organisms papaya and grape. These genomes help us understand the chromosomal history of the super-order rosids. Essential to this process are new, online genomics tools that allow researchers to easily perform their own experiments, such as identifying and evaluating syntenic regions and estimating the degree of post-tetraploidy gene fractionation (diploidization); this process is exemplified here using the online comparative genomics toolset CoGe. Using case studies, we show that two of the three genomes within the rosid paleohexaploid are more fractionated with respect to one another than to a third genome. This indicates a shared history derived from a [tetraploid]-then-[wide cross to generate a triploid]-then-[whole genome duplication to generate the hexaploid,] or similar scenario involving unreduced gametes. Two alternative hypotheses are presented that differ in terms of the mechanism and timing of fractionation.

Altered Patterns of Fractionation and Exon Deletions in Brassica Rapa Support a Two-Step Model of Paleohexaploidy

The genome sequence of the paleohexaploid Brassica rapa shows that fractionation is biased among the three subgenomes and that the least fractionated subgenome has approximately twice as many orthologs as its close (and relatively unduplicated) relative Arabidopsis than had either of the other two subgenomes. One evolutionary scenario is that the two subgenomes with heavy gene losses (I and II) were in the same nucleus for a longer period of time than the third subgenome (III) with the fewest gene losses. This “two-step” hypothesis is essentially the same as that proposed previously for the eudicot paleohexaploidy; however, the more recent nature of the B. rapa paleohexaploidy makes this model more testable. We found that subgenome II suffered recent small deletions within exons more frequently than subgenome I, as would be expected if the genes in subgenome I had already been near maximally fractionated before subgenome III was introduced. We observed that some sequences, before these deletions, were flanked by short direct repeats, a unique signature of intrachromosomal illegitimate recombination. We also found, through simulations, that short—single or two-gene—deletions appear to dominate the fractionation patterns in B. rapa. We conclude that the observed patterns of the triplicated regions in the Brassica genome are best explained by a two-step fractionation model. The triplication and subsequent mode of fractionation could influence the potential to generate morphological diversity—a hallmark of the Brassica genus.

Large-Scale Management of Insect Resistance to Transgenic Cotton in Arizona: Can Transgenic Insecticidal Crops be Sustained?

Abstract A major challenge for agriculture is management of insect resistance to toxins from Bacillus thuringiensis (Bt) produced by transgenic crops. Here we describe how a large-scale program is being developed in Arizona for management of resistance to Bt cotton in the pink bollworm, Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae), and other insect pests of cotton. Financial support from growers makes this program possible. Collaboration between the Arizona Cotton Research and Protection Council, the University of Arizona, and government agencies has led to development of resistance management guidelines, a remedial action plan, and tools for monitoring compliance with the proposed guidelines. Direct participation in development of resistance management policies is a strong incentive for growers to invest in resistance management research. However, more research, regularly updated regulations, and increased collaboration between stakeholders are urgently needed to maintain efficacy of Bt toxins in transgenic crops.

MethylCoder: software pipeline for bisulfite-treated sequences

MOTIVATION MethylCoder is a software program that generates per-base methylation data given a set of bisulfite-treated reads. It provides the option to use either of two existing short-read aligners, each with different strengths. It accounts for soft-masked alignments and overlapping paired-end reads. MethylCoder outputs data in text and binary formats in addition to the final alignment in SAM format, so that common high-throughput sequencing tools can be used on the resulting output. It is more flexible than existing software and competitive in terms of speed and memory use. AVAILABILITY MethylCoder requires only a python interpreter and a C compiler to run. Extensive documentation and the full source code are available under the MIT license at: https://github.com/brentp/methylcode. CONTACT bpederse@gmail.com.

Sources, sinks, and the zone of influence of refuges for managing insect resistance to Bt crops

The refuge strategy is central for delaying insect resistance to transgenic crops that produce Bacillus thuringiensis (Bt) toxins, but determining an effective spatial configuration of refuges has been problematic. We developed a spatially explicit, demographically based method for estimating the zone of influence of refuges, the area over which refuges increase an insects population density. The method relied on global positioning system (GPS) and geographic information system (GIS) technologies combined with spatial statistics. We applied it in two successive years to Arizona populations of pink bollworm, a major cotton pest. Refuges at 0.75 km or less from Bt cotton had the greatest potential for delaying resistance in both years studied. However, the zone of influence of refuges varied between years and among sites. The density of susceptible moths was significantly lower at sites with high compared to low abundance of Bt cotton relative to non-Bt cotton. Thus, abundance of a Bt crop relative to a non-Bt crop may influence the effectiveness of refuges. Our spatially explicit approach takes such source–sink dynamics into account and hence is well suited to help determine how refuges should be deployed.

Arabidopsis intragenomic conserved noncoding sequence

After the most recent tetraploidy in the Arabidopsis lineage, most gene pairs lost one, but not both, of their duplicates. We manually inspected the 3,179 retained gene pairs and their surrounding gene space still present in the genome using a custom-made viewer application. The display of these pairs allowed us to define intragenic conserved noncoding sequences (CNSs), identify exon annotation errors, and discover potentially new genes. Using a strict algorithm to sort high-scoring pair sequences from the bl2seq data, we created a database of 14,944 intragenomic Arabidopsis CNSs. The mean CNS length is 31 bp, ranging from 15 to 285 bp. There are ≈1.7 CNSs associated with a typical gene, and Arabidopsis CNSs are found in all areas around exons, most frequently in the 5′ upstream region. Gene ontology classifications related to transcription, regulation, or “response to …” external or endogenous stimuli, especially hormones, tend to be significantly overrepresented among genes containing a large number of CNSs, whereas protein localization, transport, and metabolism are common among genes with no CNSs. There is a 1.5% overlap between these CNSs and the 218,982 putative RNAs in the Arabidopsis Small RNA Project database, allowing for two mismatches. These CNSs provide a unique set of noncoding sequences enriched for function. CNS function is implied by evolutionary conservation and independently supported because CNS-richness predicts regulatory gene ontology categories.

G-Boxes, Bigfoot Genes, and Environmental Response: Characterization of Intragenomic Conserved Noncoding Sequences in Arabidopsis

A tetraploidy left Arabidopsis thaliana with 6358 pairs of homoeologs that, when aligned, generated 14,944 intragenomic conserved noncoding sequences (CNSs). Our previous work assembled these phylogenetic footprints into a database. We show that known transcription factor (TF) binding motifs, including the G-box, are overrepresented in these CNSs. A total of 254 genes spanning long lengths of CNS-rich chromosomes (Bigfoot) dominate this database. Therefore, we made subdatabases: one containing Bigfoot genes and the other containing genes with three to five CNSs (Smallfoot). Bigfoot genes are generally TFs that respond to signals, with their modal CNS positioned 3.1 kb 5′ from the ATG. Smallfoot genes encode components of signal transduction machinery, the cytoskeleton, or involve transcription. We queried each subdatabase with each possible 7-nucleotide sequence. Among hundreds of hits, most were purified from CNSs, and almost all of those significantly enriched in CNSs had no experimental history. The 7-mers in CNSs are not 5′- to 3′-oriented in Bigfoot genes but are often oriented in Smallfoot genes. CNSs with one G-box tend to have two G-boxes. CNSs were shared with the homoeolog only and with no other gene, suggesting that binding site turnover impedes detection. Bigfoot genes may function in adaptation to environmental change.