Survey of Candidate Genes for Maize Resistance to Infection by Aspergillus flavus and/or Aflatoxin Contamination

Many projects have identified candidate genes for resistance to aflatoxin accumulation or Aspergillus flavus infection and growth in maize using genetic mapping, genomics, transcriptomics and/or proteomics studies. However, only a small percentage of these candidates have been validated in field conditions, and their relative contribution to resistance, if any, is unknown. This study presents a consolidated list of candidate genes identified in past studies or in-house studies, with descriptive data including genetic location, gene annotation, known protein identifiers, and associated pathway information, if known. A candidate gene pipeline to test the phenotypic effect of any maize DNA sequence on aflatoxin accumulation resistance was used in this study to determine any measurable effect on polymorphisms within or linked to the candidate gene sequences, and the results are published here

Characterization of the maize lipoxygenase gene family in relation to aflatoxin accumulation resistance

<div><p>Maize (<i>Zea mays</i> L.) is a globally important staple food crop prone to contamination by aflatoxin, a carcinogenic secondary metabolite produced by the fungus <i>Aspergillus flavus</i>. An efficient approach to reduce accumulation of aflatoxin is the development of germplasm resistant to colonization and toxin production by <i>A</i>. <i>flavus</i>. Lipoxygenases (LOXs) are a group of non-heme iron containing dioxygenase enzymes that catalyze oxygenation of polyunsaturated fatty acids (PUFAs). LOX derived oxylipins play critical roles in plant defense against pathogens including <i>A</i>. <i>flavus</i>. The objectives of this study were to summarize sequence diversity and expression patterns for all LOX genes in the maize genome, and map their effect on aflatoxin accumulation via linkage and association mapping. In total, 13 LOX genes were identified, characterized, and mapped. The sequence of one gene, <i>ZmLOX10</i>, is reported from 5 inbred lines. Genes <i>ZmLOX1</i>/<i>2</i>, <i>5</i>, <i>8</i>, <i>9</i>, <i>10</i> and <i>12</i> (GRMZM2G156861, or V4 numbers ZM00001D042541 and Zm00001D042540, GRMZM2G102760, GRMZM2G104843, GRMZM2G017616, GRMZM2G015419, and GRMZM2G106748, respectively) fell under previously published QTL in one or more mapping populations and are linked to a measurable reduction of aflatoxin in maize grains. Association mapping results found 28 of the 726 SNPs tested were associated with reduced aflatoxin levels at p ≤ 9.71 x 10⁻⁴ according to association statistics. These fell within or near nine of the ZmLOX genes. This work confirms the importance of some lipoxygenases for resistance to aflatoxin accumulation and may be used to direct future genetic selection in maize.</p></div

List of SNPs, InDels, and SSRs used to map the phenotypic effect of each ZmLOXs.

<p>List of SNPs, InDels, and SSRs used to map the phenotypic effect of each ZmLOXs.</p

Sequence summary of the 5 maize inbred lines sequenced for <i>ZmLOX10</i>.

<p>Exons are filled black boxes, introns are black lines, and up-and down-stream untranslated regions are white boxes. Numbers above the gene indicate the primers used to amplify various amplicons spanning the gene (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181265#pone.0181265.s001" target="_blank">S1 Table</a>), and the ATG start site is indicated. Triangles and numbers below the gene are insertions (or deletions, if negative), and SNPs are shown in their approximate locations with the two bases indicated. Asterisks below the gene indicate SNPs that cause a non-synonymous mutation, as numbered and described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181265#pone.0181265.t005" target="_blank">Table 5</a>. Small bar to the upper right of the figure indicates the length of 100 base pairs.</p

Expression data of ZmLOX genes.

<p>Expression data of ZmLOX genes.</p

Composite interval mapping results of the MpB population (chromosome 2) for natural log transformed aflatoxin level values in one environment.

<p>The x axis represents the genetic length of the chromosome and the y axis represents the LOD. The horizontal line indicates LOD significance threshold of 2.4. GRMZM2G104843 in bin 2.04 mapped right under a QTL spanning 6.2 mbp.</p

Summary of gene association analysis for the 13 ZmLOX genes studied.

<p>Summary of gene association analysis for the 13 ZmLOX genes studied.</p

Phylogenetic tree analysis of all 13 maize LOX genes, including gene and transcript I.D. for each of the gene.

<p>The tree was constructed using the PIECE (Plant Intron and Exon Comparison Evolution) database. Green/blue color are 5’ and 3’ untranslated regions (UTRs), gray color corresponds to gene introns, and purple color to gene exons.</p

A summary of the maize lipoxygenase gene family characterized in the current study.

<p>A summary of the maize lipoxygenase gene family characterized in the current study.</p