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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.

In Saccharomyces cerevisiae, Rad51p plays a central role in homologous recombination and the repair of double-strand breaks (DSBs). Double mutants of the two Zea mays L. (maize) rad51 homologs are viable and develop well under normal conditions, but are male sterile and have substantially reduced seed set. Light microscopic analyses of male meiosis in these plants reveal reduced homologous pairing, synapsis of nonhomologous chromosomes, reduced bivalents at diakinesis, numerous chromosome breaks at anaphase I, and that >33% of quartets carry cells that either lack an organized nucleolus or have two nucleoli. This indicates that RAD51 is required for efficient chromosome pairing and its absence results in nonhomologous pairing and synapsis. These phenotypes differ from those of an Arabidopsis rad51 mutant that exhibits completely disrupted chromosome pairing and synapsis during meiosis. Unexpectedly, surviving female gametes produced by maize rad51 double mutants are euploid and exhibit near-normal rates of meiotic crossovers. The finding that maize rad51 double mutant embryos are extremely susceptible to radiation-induced DSBs demonstrates a conserved role for RAD51 in the repair of mitotic DSBs in plants, vertebrates, and yeast.

The T cytoplasm of maize serves as a model for the nuclear restoration of cytoplasmic male sterility. The rf2 gene, one of two nuclear genes required for fertility restoration in male-sterile T-cytoplasm (cmsT) maize, was cloned. The protein predicted by the rf2 sequence is a putative aldehyde dehydrogenase, which suggests several mechanisms that might explain Rf2-mediated fertility restoration in cmsT maize. Aldehyde dehydrogenase may be involved in the detoxification of acetaldehyde produced by ethanolic fermentation during pollen development, may play a role in energy metabolism, or may interact with URF13, the mitochondrial protein associated with male sterility in cmsT maize

Lateral roots are initiated from the pericycle cells of other types of roots and remain in contact with these roots throughout their life span. Although this physical contact has the potential to permit the exchange of signals, little is known about the flow of information from the lateral roots to the primary root. To begin to study these interactions the proteome of the primary root system of the maize (Zea mays L.) lrt1 mutant, which does not initiate lateral roots, was compared with the corresponding proteome of wild-type seedlings 9 days after germination. Approximately 150 soluble root proteins were resolved by two-dimensional electrophoresis and analyzed by MALDI-ToF mass spectrometry and database searching. The 96 most abundant proteins from a pH 4-7 gradient were analyzed; 67 proteins representing 47 different Genbank accessions were identified. Interestingly, 10% (15/150) of the detected proteins were preferentially expressed in lrt1 roots that lack lateral roots. Eight of these lrt1-specific proteins were identified and four are involved in lignin metabolism. This study demonstrates for the first time the influence of lateral roots on the proteome of the primary root system. To our knowledge this is the first study to demonstrate an interaction between two plant organs (viz., lateral and primary roots) at the level of the proteome.

As an ancient segmental tetraploid, the maize (Zea mays L.) genome contains large numbers of paralogs that are expected to have diverged by a minimum of 10% over time. Nearly identical paralogs (NIPs) are defined as paralogous genes that exhibit ≥98% identity. Sequence analyses of the “gene space” of the maize inbred line B73 genome, coupled with wet lab validation, have revealed that, conservatively, at least ∼1% of maize genes have a NIP, a rate substantially higher than that in Arabidopsis. In most instances, both members of maize NIP pairs are expressed and are therefore at least potentially functional. Of evolutionary significance, members of many NIP families also exhibit differential expression. The finding that some families of maize NIPs are closely linked genetically while others are genetically unlinked is consistent with multiple modes of origin. NIPs provide a mechanism for the maize genome to circumvent the inherent limitation that diploid genomes can carry at most two “alleles” per “locus.” As such, NIPs may have played important roles during the evolution and domestication of maize and may contribute to the success of long-term selection experiments in this important crop species.

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 (≈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 lowa 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 ≈33,000 and ≈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.

Rates of Mu transposon insertions and excisions are both high in late somatic cells of maize. In contrast, although high rates of insertions are observed in germinal cells, germinal excisions are recovered only rarely. Plants doubly homozygous for deletion alleles of rad51A1 and rad51A2 do not encode functional RAD51 protein (RAD51−). Approximately 1% of the gametes from RAD51+ plants that carry the MuDR-insertion allele a1-m5216 include at least partial deletions of MuDR and the a1 gene. The structures of these deletions suggest they arise via the repair of MuDR-induced double-strand breaks via nonhomologous end joining. In RAD51− plants these germinal deletions are recovered at rates that are at least 40-fold higher. These rates are not substantially affected by the presence or absence of an a1-containing homolog. Together, these findings indicate that in RAD51+ germinal cells MuDR-induced double-strand breaks (DSBs) are efficiently repaired via RAD51-directed homologous recombination with the sister chromatid. This suggests that RAD51− plants may offer an efficient means to generate deletion alleles for functional genomic studies. Additionally, the high proportion of Mu-active, RAD51− plants that exhibit severe developmental defects suggest that RAD51 plays a critical role in the repair of MuDR-induced DSBs early in vegetative development.

Recent reviews have emphasized the need to incorporate genomics into ecological field studies to further understand how species respond to changing environmental conditions. Genomic tools, such as cDNA (complementary DNA) microarrays, allow for the simultaneous analysis of gene expression of thousands of genes from all or part of an organism's genome (the transcription profile), thereby revealing the genetic mechanisms that underlie species' responses to environmental change. However, despite their potential, two major limitations have hindered the incorporation of microarrays and other genomic tools into field studies: (1) the limited availability of microarrays for ecologically relevant, non-model species and limited financial resources for developing new microarrays; and (2) concern that high sensitivity of gene expression to even subtle alterations in environmental conditions will hinder detection of relevant changes in field measures of transcription profiles. Here, we show that with cross-species hybridizations of microarrays developed for a closely related model organism, an appropriate experimental design, and sufficient replication, transcriptional profiling can successfully be incorporated into field studies. In this way, relevant changes in gene expression with changing environmental conditions can be detected.

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.

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-C0t (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.