Biomedical subjects

Patrick S Schnable

Publications and source records attributed to Patrick S Schnable.

At least 19 recordsLinked to original sources

PubMed2006

Genetic dissection of intermated recombinant inbred lines using a new genetic map of maize.

A new genetic map of maize, ISU-IBM Map4, that integrates 2029 existing markers with 1329 new indel polymorphism (IDP) markers has been developed using intermated recombinant inbred lines (IRILs) from the intermated B73xMo17 (IBM) population. The website http://magi.plantgenomics.iastate.edu provides access to IDP primer sequences, sequences from which IDP primers were designed, optimized marker-specific PCR conditions, and polymorphism data for all IDP markers. This new gene-based genetic map will facilitate a wide variety of genetic and genomic research projects, including map-based genome sequencing and gene cloning. The mosaic structures of the genomes of 91 IRILs, an important resource for identifying and mapping QTL and eQTL, were defined. Analyses of segregation data associated with markers genotyped in three B73/Mo17-derived mapping populations (F2, Syn5, and IBM) demonstrate that allele frequencies were significantly altered during the development of the IBM IRILs. The observations that two segregation distortion regions overlap with maize flowering-time QTL suggest that the altered allele frequencies were a consequence of inadvertent selection. Detection of two-locus gamete disequilibrium provides another means to extract functional genomic data from well-characterized plant RILs.

Alleles↗

PubMed2006

Effects of trans-acting genetic modifiers on meiotic recombination across the a1-sh2 interval of maize.

Meiotic recombination rates are potentially affected by cis- and trans-acting factors, i.e., genotype-specific modifiers that do or do not reside in the recombining interval, respectively. Effects of trans modifiers on recombination across the approximately 140-kb maize a1-sh2 interval of chromosome 3L were studied in the absence of polymorphic cis factors in three genetically diverse backgrounds into which a sequence-identical a1-sh2 interval had been introgressed. Genetic distances across a1-sh2 varied twofold among genetic backgrounds. Although the existence of regions exhibiting high and low rates of recombination (hot and cold spots, respectively) was conserved across backgrounds, the absolute rates of recombination in these sequence-identical regions differed significantly among backgrounds. In addition, an intergenic hot spot had a higher rate of recombination as compared to the genome average rate of recombination in one background and not in another. Recombination rates across two genetic intervals on chromosome 1 did not exhibit the same relationships among backgrounds as was observed in a1-sh2. This suggests that at least some detected trans-acting factors do not equally affect recombination across the genome. This study establishes that trans modifier(s) polymorphic among genetic backgrounds can increase and decrease recombination in both genic and intergenic regions over relatively small genetic and physical intervals.

Chromosomal Instability↗

PubMed2006

Unequal sister chromatid and homolog recombination at a tandem duplication of the A1 locus in maize.

Tandemly arrayed duplicate genes are prevalent. The maize A1-b haplotype is a tandem duplication that consists of the components, alpha and beta. The rate of meiotic unequal recombination at A1-b is ninefold higher when a homolog is present than when it is absent (i.e., hemizygote). When a sequence heterologous homolog is available, 94% of recombinants (264/281) are generated via recombination with the homolog rather than with the sister chromatid. In addition, 83% (220/264) of homolog recombination events involved alpha rather than beta. These results indicate that: (1) the homolog is the preferred template for unequal recombination and (2) pairing of the duplicated segments with the homolog does not occur randomly but instead favors a particular configuration. The choice of recombination template (i.e., homolog vs. sister chromatid) affects the distribution of recombination breakpoints within a1. Rates of unequal recombination at A1-b are similar to the rate of recombination between nonduplicated a1 alleles. Unequal recombination is therefore common and is likely to be responsible for the generation of genetic variability, even within inbred lines.

Base Sequence↗

PubMed2006

Scanning microarrays at multiple intensities enhances discovery of differentially expressed genes.

MOTIVATION: Scanning parameters are often overlooked when optimizing microarray experiments. A scanning approach that extends the dynamic data range by acquiring multiple scans of different intensities has been developed. RESULTS: Data from each of three scan intensities (low, medium, high) were analyzed separately using multiple scan and linear regression approaches to identify and compare the sets of genes that exhibit statistically significant differential expression. In the multiple scan approach only one-third of the differentially expressed genes were shared among the three intensities, and each scan intensity identified unique sets of differentially expressed genes. The set of differentially expressed genes from any one scan amounted to < 70% of the total number of genes identified in at least one scan. The average signal intensity of genes that exhibited statistically significant changes in expression was highest for the low-intensity scan and lowest for the high-intensity scan, suggesting that low-intensity scans may be best for detecting expression differences in high-signal genes, while high-intensity scans may be best for detecting expression differences in low-signal genes. Comparison of the differentially expressed genes identified in the multiple scan and linear regression approaches revealed that the multiple scan approach effectively identifies a subset of statistically significant genes that linear regression approach is unable to identify. Quantitative RT-PCR (qRT-PCR) tests demonstrated that statistically significant differences identified at all three scan intensities can be verified. AVAILABILITY: The data presented can be viewed at http://www.ncbi.nlm.nih.gov/geo/ under GEO accession no. GSE3017.

Gene Expression Profiling↗

PubMed2006

All possible modes of gene action are observed in a global comparison of gene expression in a maize F1 hybrid and its inbred parents.

Heterosis is the phenomenon whereby the progeny of particular inbred lines have enhanced agronomic performance relative to both parents. Although several hypotheses have been proposed to explain this fundamental biological phenomenon, the responsible molecular mechanisms have not been determined. The maize inbred lines B73 and Mo17 produce a heterotic F1 hybrid. Global patterns of gene expression were compared in seedlings of these three genotypes by using a microarray that contains 13,999 cDNAs. Using an estimated 15% false discovery rate as a cutoff, 1,367 ESTs (9.8%) were identified as being significantly differentially expressed among genotypes. All possible modes of gene action were observed, including additivity, high- and low-parent dominance, underdominance, and overdominance. The largest proportion of the ESTs (78%; 1,062 of 1,367) exhibited expression patterns that are not statistically distinguishable from additivity. Even so, 22% of the differentially regulated ESTs exhibited nonadditive modes of gene expression. Classified on the basis of significant pairwise comparisons of genotype means, 181 of these 305 nonadditive ESTs exhibited high-parent dominance and 23 exhibited low-parent dominance. In addition, 44 ESTs exhibited underdominance or overdominance. These findings are consistent with the hypothesis that multiple molecular mechanisms, including overdominance, contribute to heterosis.

Expressed Sequence Tags↗

PubMed2006

GRAMA: genetic mapping analysis of temperature gradient capillary electrophoresis data.

Temperature gradient capillary electrophoresis (TGCE) is a high-throughput method to detect segregating single nucleotide polymorphisms and InDel polymorphisms in genetic mapping populations. Existing software that analyzes TGCE data was, however, designed for mutation analysis rather than genetic mapping. Genetic recombinant analysis and mapping assistant (GRAMA) is a new tool that automates TGCE data analysis for the purpose of genetic mapping. Data from multiple TGCE runs are analyzed, integrated, and displayed in an intuitive visual format. GRAMA includes an algorithm to detect peaks in electropherograms and can automatically compare its peak calls with those produced by another software package. Consequently, GRAMA provides highly accurate results with a low false positive rate of 5.9% and an even lower false negative rate of 1.3%. Because of its accuracy and intuitive interface, GRAMA boosts user productivity more than twofold relative to previous manual methods of scoring TGCE data. GRAMA is written in Java and is freely available at http://www.complex.iastate.edu .

Algorithms↗

PubMed2005

Isolation, characterization, and pericycle-specific transcriptome analyses of the novel maize lateral and seminal root initiation mutant rum1.

The monogenic recessive maize (Zea mays) mutant rootless with undetectable meristems 1 (rum1) is deficient in the initiation of the embryonic seminal roots and the postembryonic lateral roots at the primary root. Lateral root initiation at the shoot-borne roots and development of the aerial parts of the mutant rum1 are not affected. The mutant rum1 displays severely reduced auxin transport in the primary root and a delayed gravitropic response. Exogenously applied auxin does not induce lateral roots in the primary root of rum1. Lateral roots are initiated in a specific cell type, the pericycle. Cell-type-specific transcriptome profiling of the primary root pericycle 64 h after germination, thus before lateral root initiation, via a combination of laser capture microdissection and subsequent microarray analyses of 12k maize microarray chips revealed 90 genes preferentially expressed in the wild-type pericycle and 73 genes preferentially expressed in the rum1 pericycle (fold change >2; P-value <0.01; estimated false discovery rate of 13.8%). Among the 51 annotated genes predominately expressed in the wild-type pericycle, 19 genes are involved in signal transduction, transcription, and the cell cycle. This analysis defines an array of genes that is active before lateral root initiation and will contribute to the identification of checkpoints involved in lateral root formation downstream of rum1.

Biological Transport↗

PubMed2005

Quality assessment of maize assembled genomic islands (MAGIs) and large-scale experimental verification of predicted genes.

Recent sequencing efforts have targeted the gene-rich regions of the maize (Zea mays L.) genome. We report the release of an improved assembly of maize assembled genomic islands (MAGIs). The 114,173 resulting contigs have been subjected to computational and physical quality assessments. Comparisons to the sequences of maize bacterial artificial chromosomes suggest that at least 97% (160 of 165) of MAGIs are correctly assembled. Because the rates at which junction-testing PCR primers for genomic survey sequences (90-92%) amplify genomic DNA are not significantly different from those of control primers ( approximately 91%), we conclude that a very high percentage of genic MAGIs accurately reflect the structure of the maize genome. EST alignments, ab initio gene prediction, and sequence similarity searches of the MAGIs are available at the Iowa State University MAGI web site. This assembly contains 46,688 ab initio predicted genes. The expression of almost half (628 of 1,369) of a sample of the predicted genes that lack expression evidence was validated by RT-PCR. Our analyses suggest that the maize genome contains between approximately 33,000 and approximately 54,000 expressed genes. Approximately 5% (32 of 628) of the maize transcripts discovered do not have detectable paralogs among maize ESTs or detectable homologs from other species in the GenBank NR nucleotide/protein database. Analyses therefore suggest that this assembly of the maize genome contains approximately 350 previously uncharacterized expressed genes. We hypothesize that these "orphans" evolved quickly during maize evolution and/or domestication.

Chromosomes, Artificial, Bacterial↗

PubMed2005

The roothairless1 gene of maize encodes a homolog of sec3, which is involved in polar exocytosis.

The roothairless1 (rth1) mutant is impaired in root hair elongation and exhibits other growth abnormalities. Unicellular root hairs elongate via localized tip growth, a process mediated by polar exocytosis of secretory vesicles. We report here the cloning of the rth1 gene that encodes a sec3 homolog. In yeast (Saccharomyces cerevisiae) and mammals, sec3 is a subunit of the exocyst complex, which tethers exocytotic vesicles prior to their fusion. The cloning of the rth1 gene associates the homologs of exocyst subunits to an exocytotic process in plant development and supports the hypothesis that exocyst-like proteins are involved in plant exocytosis. Proteomic analyses identified four proteins that accumulate to different levels in wild-type and rth1 primary roots. The preferential accumulation in the rth1 mutant proteome of a negative regulator of the cell cycle (a prohibitin) may at least partially explain the delayed development and flowering of the rth1 mutant.

Chromosome Mapping↗

PubMed2005

Cis-effects on meiotic recombination across distinct a1-sh2 intervals in a common Zea genetic background.

Genetic distances across the a1-sh2 interval varied threefold in three near-isogenic stocks that carry structurally distinct teosinte A1 Sh2 haplotypes (from Z. mays spp. mexicana Chalco, Z. mays spp. parviglumis, and Z. luxurians) and a common maize a1::rdt sh2 haplotype. In each haplotype >85% of recombination events resolved in the proximal 10% of the approximately 130-kb a1-sh2 interval. Even so, significant differences in the distributions of recombination breakpoints were observed across subintervals among haplotypes. Each of the three previously detected recombination hot spots was detected in at least one of the three teosinte haplotypes and two of these hot spots were not detected in at least one teosinte haplotype. Moreover, novel hot spots were detected in two teosinte haplotypes. Due to the near-isogenic nature of the three stocks, the observed variation in the distribution of recombination events is the consequence of cis-modifications. Although generally negatively correlated with rates of recombination per megabase, levels of sequence polymorphisms do not fully account for the nonrandom distribution of recombination breakpoints. This study also suggests that estimates of linkage disequilibrium must be interpreted with caution when considering whether a gene has been under selection.

Chi-Square Distribution↗

PubMed2005

Temperature gradient capillary electrophoresis (TGCE)--a tool for the high-throughput discovery and mapping of SNPs and IDPs.

Temperature gradient capillary electrophoresis (TGCE) can be used to distinguish heteroduplex from homoduplex DNA molecules and can thus be applied to the detection of various types of DNA polymorphisms. Unlike most single nucleotide polymorphism (SNP) detection technologies, TGCE can be used even in the absence of prior knowledge of the sequences of the underlying polymorphisms. TGCE is both sensitive and reliable in detecting SNPs, small InDel (insertion/deletion) polymorphisms (IDPs) and simple sequence repeats, and using this technique it is possible to detect a single SNP in amplicons of over 800 bp and 1-bp IDPs in amplicons of approximately 500 bp. Genotyping data obtained via TGCE are consistent with data obtained via gel-based detection technologies. For genetic mapping experiments, TGCE has a number of advantages over alternative heteroduplex-detection technologies such as celery endonuclease (CELI) and denaturing high-performance liquid chromatography (dHPLC). Multiplexing can increase TGCE's throughput to 12 markers on 94 recombinant inbreds per day. Given its ability to efficiently and reliably detect a variety of subtle DNA polymorphisms that occur at high frequency in genes, TGCE shows great promise for discovering polymorphisms and conducting genetic mapping and genotyping experiments.

Chromosome Mapping↗

PubMed2005

The accumulation of abundant soluble proteins changes early in the development of the primary roots of maize (Zea mays L.).

A reference database of the major soluble proteins of the primary root of the maize inbred line B73 was generated 5 days after germination (DAG) using a combination of 2-DE and MALDI-TOF MS. A total of 302 protein spots were detected with CBB in a pH 4-7 range and 81 proteins representing 74 distinct Genbank accessions were identified. Only 28% of the major proteins identified in 5 DAG primary roots were identified in similarly analyzed 9 DAG primary roots documenting remarkable changes in the accumulation of abundant soluble proteins early in primary root development.

Electrophoresis, Gel, Two-Dimensional↗

PubMed2005

Evaluation of five ab initio gene prediction programs for the discovery of maize genes.

Five ab initio programs (FGENESH, GeneMark.hmm, GENSCAN, GlimmerR and Grail) were evaluated for their accuracy in predicting maize genes. Two of these programs, GeneMark.hmm and GENSCAN had been trained for maize; FGENESH had been trained for monocots (including maize), and the others had been trained for rice or Arabidopsis. Initial evaluations were conducted using eight maize genes (gl8a, pdc2, pdc3, rf2c, rf2d, rf2e1, rth1, and rth3) of which the sequences were not released to the public prior to conducting this evaluation. The significant advantage of this data set for this evaluation is that these genes could not have been included in the training sets of the prediction programs. FGENESH yielded the most accurate and GeneMark.hmm the second most accurate predictions. The five programs were used in conjunction with RT-PCR to identify and establish the structures of two new genes in the a1-sh2 interval of the maize genome. FGENESH, GeneMark.hmm and GENSCAN were tested on a larger data set consisting of maize assembled genomic islands (MAGIs) that had been aligned to ESTs. FGENESH, GeneMark.hmm and GENSCAN correctly predicted gene models in 773, 625, and 371 MAGIs, respectively, out of the 1353 MAGIs that comprise data set 2.

Alternative Splicing↗

PubMed2005

Consensus genetic maps: a graph theoretic approach.

A genetic map is an ordering of genetic markers constructed from genetic linkage data for use in linkage studies and experimental design. While traditional methods have focused on constructing maps from a single population study, increasingly maps are generated for multiple lines and populations of the same organism. For example, in crop plants, where the genetic variability is high, researchers have created maps for many populations. In the face of these new data, we address the increasingly important problem of generating a consensus map - an ordering of all markers in the various population studies. In our method, each input map is treated as a partial order on a set of markers. To find the most consistent order shared between maps, we model the partial orders as directed graphs. We create an aggregate by merginging the transitive closure of the input graphs and taking the transitive reduction of the result. In this process, cycles may need to be broken to resolve inconsistencies between the inputs. The cycle breaking problem is NP-hard, but the problem size depends upon the scope of the inconsistency between the input graphs, which will be local if the input graphs are from closely related organisms. We present results of running the resulting software on maps generated from seven populations of the crop plant Zea Mays.

Chromosome Mapping↗

PubMed2005

Characterization of two GL8 paralogs reveals that the 3-ketoacyl reductase component of fatty acid elongase is essential for maize (Zea mays L.) development.

Prior analyses established that the maize (Zea mays L.) gl8a gene encodes 3-ketoacyl reductase, a component of the fatty acid elongase required for the biosynthesis of very long chain fatty acids (VLCFAs). A paralogous gene, gl8b, has been identified that is 96% identical to gl8a. The gl8a and gl8b genes map to syntenic chromosomal regions, have similar, but not identical, expression patterns, and encode proteins that are 97% identical. Both of these genes are required for the normal accumulation of cuticular waxes on seedling leaves. The chemical composition of the cuticular waxes from gl8a and gl8b mutants indicates that these genes have at least overlapping, if not redundant, functions in cuticular wax biosynthesis. Although gl8a and gl8b double mutant kernels have endosperms that cannot be distinguished from wild-type siblings, these kernels are non-viable because their embryos fail to undergo normal development. Double mutant kernels accumulate substantially reduced levels of VLCFAs. VLCFAs are components of a variety of compounds, for example, cuticular waxes, suberin, and sphingolipids. Consistent with their essential nature in yeast, the accumulation of the ceramide moiety of sphingolipids is substantially reduced and their fatty acid composition altered in gl8a and gl8b double mutant kernels relative to wild-type kernels. Hence, we hypothesize that sphingolipids or other VLCFA-containing compounds are essential for normal embryo development.

3-Oxoacyl-(Acyl-Carrier-Protein) Reductase↗

PubMed2004

MuDR transposase increases the frequency of meiotic crossovers in the vicinity of a Mu insertion in the maize a1 gene.

Although DNA breaks stimulate mitotic recombination in plants, their effects on meiotic recombination are not known. Recombination across a maize a1 allele containing a nonautonomous Mu transposon was studied in the presence and absence of the MuDR-encoded transposase. Recombinant A1' alleles isolated from a1-mum2/a1::rdt heterozygotes arose via either crossovers (32 CO events) or noncrossovers (8 NCO events). In the presence of MuDR, the rate of COs increased fourfold. This increase is most likely a consequence of the repair of MuDR-induced DNA breaks at the Mu1 insertion in a1-mum2. Hence, this study provides the first in vivo evidence that DNA breaks stimulate meiotic crossovers in plants. The distribution of recombination breakpoints is not affected by the presence of MuDR in that 19 of 24 breakpoints isolated from plants that carried MuDR mapped to a previously defined 377-bp recombination hotspot. This result is consistent with the hypothesis that the DNA breaks that initiate recombination at a1 cluster at its 5' end. Conversion tracts associated with eight NCO events ranged in size from <700 bp to >1600 bp. This study also establishes that MuDR functions during meiosis and that ratios of CO/NCO vary among genes and can be influenced by genetic background.

Alleles↗

PubMed2004

Types and frequencies of sequencing errors in methyl-filtered and high c0t maize genome survey sequences.

The Maize Genome Sequencing Consortium has deposited into GenBank more than 850,000 maize (Zea mays) genome survey sequences (GSSs) generated via two gene enrichment strategies, methylation filtration and high-C(0)t (HC) fractionation. These GSSs are a valuable resource for generating genome assemblies and the discovery of single nucleotide polymorphisms and nearly identical paralogs. Based on the rate of mismatches between 183 GSSs (105 methylation filtration + 78 HC) and 10 control genes, the rate of sequencing errors in these GSSs is 2.3 x 10(-3). As expected many of these errors were derived from insufficient vector trimming and base-calling errors. Surprisingly, however, some errors were due to cloning artifacts. These G.C to A.T transitions are restricted to HC clones; over 40% of HC clones contain at least one such artifact. Because it is not possible to distinguish the cloning artifacts from biologically relevant polymorphisms, HC sequences should be used with caution for the discovery of single nucleotide polymorphisms or paramorphisms. The average rate of sequencing errors was reduced 6-fold (to 3.6 x 10(-4)) by applying more stringent trimming parameters. This trimming resulted in the loss of only 11% of the bases (15,469/144,968). Due to redundancy among GSSs this more stringent trimming reduced coverage of promoters, exons, and introns by only 0%, 1%, and 4%, respectively. Hence, at the cost of a very modest loss of gene coverage, the quality of these maize GSSs can approach Bermuda standards, even prior to assembly.

DNA, Plant↗

PubMed2004

Picky: oligo microarray design for large genomes.

MOTIVATION: Many large genomes are getting sequenced nowadays. Biologists are eager to start microarray analysis taking advantage of all known genes of a species, but existing microarray design tools were very inefficient for large genomes. Also, many existing tools operate in a batch mode that does not assure best designs. RESULTS: Picky is an efficient oligo microarray design tool for large genomes. Picky integrates novel computer science techniques and the best known nearest-neighbor parameters to quickly identify sequence similarities and estimate their hybridization properties. Oligos designed by Picky are computationally optimized to guarantee the best specificity, sensitivity and uniformity under the given design constrains. Picky can be used to design arrays for whole genomes, or for only a subset of genes. The latter can still be screened against a whole genome to attain the same quality as a whole genome array, thereby permitting low budget, pathway-specific experiments to be conducted with large genomes. Picky is the fastest oligo array design tool currently available to the public, requiring only a few hours to process large gene sets from rice, maize or human.

Algorithms↗