Genome assembly and visualization of aggressive wheat blast strain 16MoT01

Wheat blast, a highly destructive fungal disease caused by pathotype Magnaporthe oryzae Triticum, can cause up to 100% yield loss in wheat fields under optimal pathogen conditions. Until 2016, the disease had been confined to South America, but recent outbreaks of the disease in Asia and Africa threaten the global wheat supply. This study aims to characterize the genome structure of strain 16MoT01, which has proven to be particularly aggressive even towards wheat genotypes that have previously been resistant to blast. Genomic DNA from 16MoT01 was sequenced using Oxford Nanopore long read sequencing, assembled with Canu, and polished using Illumina reads, resulting in a finished chromosome-level assembly consisting of seven core-chromosomes, a mini-chromosome, and a mitochondrial genome. When compared to the reference genome of strain B71, the core-chromosomes show high similarity and the mini-chromosome shows a high level of divergence. The presence of mini-chromosomes will be confirmed through contour-clamped electric field (CHEF) gel electrophoresis. The CHEF protocol was developed using genomic DNA from a rice blast fungus. This assembly provides another reference genome and potential insights into what makes this strain so aggressive

HB-PLS: A statistical method for identifying biological process or pathway regulators by integrating Huber loss and Berhu penalty with partial least squares regression

Gene expression data features high dimensionality, multicollinearity, and non-Gaussian distribution noise, posing hurdles for identification of true regulatory genes controlling a biological process or pathway. In this study, we integrated the Huber loss function and the Berhu penalty (HB) into partial least squares (PLS) framework to deal with the high dimension and multicollinearity property of gene expression data, and developed a new method called HB-PLS regression to model the relationships between regulatory genes and pathway genes. To solve the Huber-Berhu optimization problem, an accelerated proximal gradient descent algorithm with at least 10 times faster than the general convex optimization solver (CVX), was developed. Application of HB-PLS to recognize pathway regulators of lignin biosynthesis and photosynthesis in Arabidopsis thaliana led to the identification of many known positive pathway regulators that had previously been experimentally validated. As compared to sparse partial least squares (SPLS) regression, an efficient method for variable selection and dimension reduction in handling multicollinearity, HB-PLS has higher efficacy in identifying more positive known regulators, a much higher but slightly less sensitivity/(1-specificity) in ranking the true positive known regulators to the top of the output regulatory gene lists for the two aforementioned pathways. In addition, each method could identify some unique regulators that cannot be identified by the other methods. Our results showed that the overall performance of HB-PLS slightly exceeds that of SPLS but both methods are instrumental for identifying real pathway regulators from high-throughput gene expression data, suggesting that integration of statistics, machine leaning and convex optimization can result in a method with high efficacy and is worth further exploration

Regulation of regeneration in Arabidopsis thaliana

We employed several algorithms with high efficacy to analyze the public transcriptomic data, aiming to identify key transcription factors (TFs) that regulate regeneration in Arabidopsis thaliana. Initially, we utilized CollaborativeNet, also known as TF-Cluster, to construct a collaborative network of all TFs, which was subsequently decomposed into many subnetworks using the Triple-Link and Compound Spring Embedder (CoSE) algorithms. Functional analysis of these subnetworks led to the identification of nine subnetworks closely associated with regeneration. We further applied principal component analysis and gene ontology (GO) enrichment analysis to reduce the subnetworks from nine to three, namely subnetworks 1, 12, and 17. Searching for TF-binding sites in the promoters of the co-expressed and co-regulated (CCGs) genes of all TFs in these three subnetworks and Triple-Gene Mutual Interaction analysis of TFs in these three subnetworks with the CCGs involved in regeneration enabled us to rank the TFs in each subnetwork. Finally, six potential candidate TFs—WOX9A, LEC2, PGA37, WIP5, PEI1, and AIL1 from subnetwork 1—were identified, and their roles in somatic embryogenesis (GO:0010262) and regeneration (GO:0031099) were discussed, so were the TFs in Subnetwork 12 and 17 associated with regeneration. The TFs identified were also assessed using the CIS-BP database and Expression Atlas. Our analyses suggest some novel TFs that may have regulatory roles in regeneration and embryogenesis and provide valuable data and insights into the regulatory mechanisms related to regeneration. The tools and the procedures used here are instrumental for analyzing high-throughput transcriptomic data and advancing our understanding of the regulation of various biological processes of interest

Emergence of a New Population of Rathayibacter toxicus: An Ecologically Complex, Geographically Isolated Bacterium

Citation: Arif, M., Busot, G. Y., Mann, R., Rodoni, B., Liu, S. Z., & Stack, J. P. (2016). Emergence of a New Population of Rathayibacter toxicus: An Ecologically Complex, Geographically Isolated Bacterium. Plos One, 11(5), 20. https://doi.org/10.1371/journal.pone.0156182Rathayibacter toxicus is a gram-positive bacterium that infects the floral parts of several Poaceae species in Australia. Bacterial ooze is often produced on the surface of infected plants and bacterial galls are produced in place of seed. R. toxicus is a regulated plant pathogen in the U.S. yet reliable detection and diagnostic tools are lacking. To better understand this geographically-isolated plant pathogen, genetic variation as a function of geographic location, host species, and date of isolation was determined for isolates collected over a forty-year period. Discriminant analyses of recently collected and archived isolates using Multi-Locus Sequence Typing (MLST) and Inter-Simple Sequence Repeats (ISSR) identified three populations of R. toxicus; RT-I and RT-II from South Australia and RT-III from Western Australia. Population RT-I, detected in 2013 and 2014 from the Yorke Peninsula in South Australia, is a newly emerged population of R. toxicus not previously reported. Commonly used housekeeping genes failed to discriminate among the R. toxicus isolates. However, strategically selected and genome-dispersed MLST genes representing an array of cellular functions from chromosome replication, antibiotic resistance and biosynthetic pathways to bacterial acquired immunity were discriminative. Genetic variation among isolates within the RT-I population was less than the within-population variation for the previously reported RT-II and RT-III populations. The lower relative genetic variation within the RT-I population and its absence from sampling over the past 40 years suggest its recent emergence. RT-I was the dominant population on the Yorke Peninsula during the 2013-2014 sampling period perhaps indicating a competitive advantage over the previously detected RT-II population. The potential for introduction of this bacterial plant pathogen into new geographic areas provide a rationale for understanding the ecological and evolutionary trajectories of R. toxicus

Massive Shift in Gene Expression during Transitions between Developmental Stages of the Gall Midge, Mayetiola Destructor

Citation: Chen, M. S., Liu, S. Z., Wang, H. Y., Cheng, X. Y., El Bouhssini, M., & Whitworth, R. J. (2016). Massive Shift in Gene Expression during Transitions between Developmental Stages of the Gall Midge, Mayetiola Destructor. PLoS One, 11(5), 1-19. https://doi.org/10.1371/journal.pone.0155616Mayetiola destructor is a destructive pest of wheat and has six developmental stages. Molecular mechanisms controlling the transition between developmental stages remain unknown. Here we analyzed genes that were expressed differentially between two successive developmental stages, including larvae at 1, 3, 5, and 7 days, pupae, and adults. A total of 17,344 genes were expressed during one or more of these studied stages. Among the expressed genes, 38-68% were differently expressed between two successive stages, with roughly equal percentages of up-and down-regulated genes. Analysis of the functions of the differentially expressed genes revealed that each developmental stage had some unique types of expressed genes that are characteristic of the physiology at that stage. This is the first genome-wide analysis of genes differentially expressed in different stages in a gall midge. The large dataset of up-and down-regulated genes in each stage of the insect shall be very useful for future research to elucidate mechanisms regulating insect development and other biological processes

Complete genome sequence of the African strain AXO1947 of Xanthomonas oryzae pv. oryzae

Citation: Huguet-Tapia, J. C., Peng, Z., Yang, B., Yin, Z., Liu, S., & White, F. F. (2016). Complete genome sequence of the African strain AXO1947 of Xanthomonas oryzae pv. oryzae. Genome Announcements, 4(1). doi:10.1128/genomeA.01730-15Citation: Tapia, J., . . . & White, F. (2013). Complete Genome Sequence of the African Strain AXO1947 of Xanthomonas oryzae pv. oryzae. Genome Announcements, 4(1). https://doi.org/10.1128/genomeA.01730-15Xanthomonas oryzae pv. oryzae is the etiological agent of bacterial rice blight. Three distinct clades of X. oryzae pv. oryzae are known. We present the complete annotated genome of the African clade strain AXO194 using long-read single-molecule PacBio sequencing technology. The genome comprises a single chromosome of 4,674,975 bp and encodes for nine transcriptional activator-like (TAL) effectors. The approach and data presented in this announcement provide information for complex bacterial genome organization and the discovery of new virulence effectors, and they facilitate target characterization of TAL effectors. © 2016 Huguet-Tapia et al

Genes Expressed Differentially in Hessian Fly Larvae Feeding in Resistant and Susceptible Plants

Citation: Chen, M. S., Liu, S. Z., Wang, H. Y., Cheng, X. Y., El Bouhssini, M., & Whitworth, R. J. (2016). Genes Expressed Differentially in Hessian Fly Larvae Feeding in Resistant and Susceptible Plants. International Journal of Molecular Sciences, 17(8), 14. https://doi.org/10.3390/ijms17081324The Hessian fly, Mayetiola destructor, is a destructive pest of wheat worldwide and mainly controlled by deploying resistant cultivars. In this study, we investigated the genes that were expressed differentially between larvae in resistant plants and those in susceptible plants through RNA sequencing on the Illumina platform. Informative genes were 11,832, 14,861, 15,708, and 15,071 for the comparisons between larvae in resistant versus susceptible plants for 0.5, 1, 3, and 5 days, respectively, after larvae had reached the feeding site. The transcript abundance corresponding to 5401, 6902, 8457, and 5202 of the informative genes exhibited significant differences (p <= 0.05) in the respective paired comparisons. Overall, genes involved in nutrient metabolism, RNA and protein synthesis exhibited lower transcript abundance in larvae from resistant plants, indicating that resistant plants inhibited nutrient metabolism and protein production in larvae. Interestingly, the numbers of cytochrome P450 genes with higher transcript abundance in larvae from resistant plants were comparable to, or higher than those with lower transcript abundance, indicating that toxic chemicals from resistant plants may have played important roles in Hessian fly larval death. Our study also identified several families of genes encoding secreted salivary gland proteins (SSGPs) that were expressed at early stage of 1st instar larvae and with more genes with higher transcript abundance in larvae from resistant plants. Those SSGPs are candidate effectors with important roles in plant manipulation

tGBS® genotyping-by-sequencing enables reliable genotyping of heterozygous loci

Conventional genotyping-by-sequencing (cGBS) strategies suffer from high rates of missing data and genotyping errors, particularly at heterozygous sites. tGBS® genotyping-by-sequencing is a novel method of genome reduction that employs two restriction enzymes to generate overhangs in opposite orientations to which (single-strand) oligos rather than (double-stranded) adaptors are ligated. This strategy ensures that only doubledigested fragments are amplified and sequenced. The use of oligos avoids the necessity of preparing adaptors and the problems associated with inter-adaptor annealing/ligation. Hence, the tGBS protocol simplifies the preparation of high-quality GBS sequencing libraries. During polymerase chain reaction (PCR) amplification, selective nucleotides included at the 3\u27-end of the PCR primers result in additional genome reduction as compared to cGBS. By adjusting the number of selective bases, different numbers of genomic sites are targeted for sequencing. Therefore, for equivalent amounts of sequencing, more reads per site are available for SNP calling. Hence, as compared to cGBS, tGBS delivers higher SNP calling accuracy (\u3e97–99%), even at heterozygous sites, less missing data per marker across a population of samples, and an enhanced ability to genotype rare alleles. tGBS is particularly well suited for genomic selection, which often requires the ability to genotype populations of individuals that are heterozygous at many loci

A Comprehensive Analysis of Alternative Splicing in Paleopolyploid Maize

Citation: Mei, W. B., Liu, S. Z., Schnable, J. C., Yeh, C. T., Springer, N. M., Schnable, P. S., & Barbazuk, W. B. (2017). A Comprehensive Analysis of Alternative Splicing in Paleopolyploid Maize. Frontiers in Plant Science, 8, 19. https://doi.org/10.3389/fpls.2017.00694Identifying and characterizing alternative splicing (AS) enables our understanding of the biological role of transcript isoform diversity. This study describes the use of publicly available RNA-Seq data to identify and characterize the global diversity of AS isoforms in maize using the inbred lines B73 and Mo17, and a related species, sorghum. Identification and characterization of AS within maize tissues revealed that genes expressed in seed exhibit the largest differential AS relative to other tissues examined. Additionally, differences in AS between the two genotypes B73 and Mo17 are greatest within genes expressed in seed. We demonstrate that changes in the level of alternatively spliced transcripts (intron retention and exon skipping) do not solely reflect differences in total transcript abundance, and we present evidence that intron retention may act to fine-tune gene expression across seed development stages. Furthermore, we have identified temperature sensitive AS in maize and demonstrate that drought-induced changes in AS involve distinct sets of genes in reproductive and vegetative tissues. Examining our identified AS isoforms within B73 X Mo17 recombinant inbred lines (RILs) identified splicing QTL (sQTL). The 43.3% of cis-sQTL regulated junctions are actually identified as alternatively spliced junctions in our analysis, while 10 Mb windows on each side of 48.2% of trans-sQTLs overlap with splicing related genes. Using sorghum as an out-group enabled direct examination of loss or conservation of AS between homeologous genes representing the two subgenomes of maize. We identify several instances where AS isoforms that are conserved between one maize homeolog and its sorghum ortholog are absent from the second maize homeolog, suggesting that these AS isoforms may have been lost after the maize whole genome duplication event. This comprehensive analysis provides new insights into the complexity of AS in maize

Characterization of maize roothairless6 which encodes a D-type cellulose synthase and controls the switch from bulge formation to tip growth

Citation: Li, L., Hey, S., Liu, S. Z., Liu, Q., McNinch, C., Hu, H. C., . . . Hochholdinger, F. (2016). Characterization of maize roothairless6 which encodes a D-type cellulose synthase and controls the switch from bulge formation to tip growth. Scientific Reports, 6, 12. doi:10.1038/srep34395Root hairs are tubular extensions of the epidermis. Root hairs of the monogenic recessive maize mutant roothairless 6 (rth6) are arrested after bulge formation during the transition to tip growth and display a rough cell surface. BSR-Seq in combination with Seq-walking and subsequent analyses of four independently generated mutant alleles established that rth6 encodes CSLD5 a plasma membrane localized 129 kD D-type cellulose synthase with eight transmembrane domains. Cellulose synthases are required for the biosynthesis of cellulose, the most abundant biopolymer of plant cell walls. Phylogenetic analyses revealed that RTH6 is part of a monocot specific clade of D-type cellulose synthases. D-type cellulose synthases are highly conserved in the plant kingdom with five gene family members in maize and homologs even among early land plants such as the moss Physcomitrella patens or the clubmoss Selaginella moellendorffii. Expression profiling demonstrated that rth6 transcripts are highly enriched in root hairs as compared to all other root tissues. Moreover, in addition to the strong knock down of rth6 expression in young primary roots of the mutant rth6, the gene is also significantly down-regulated in rth3 and rth5 mutants, while it is up-regulated in rth2 mutants, suggesting that these genes interact in cell wall biosynthesis