BackgroundDrought stress is one of the major limiting factors for maize production. With the availability of maize B73 reference genome and whole-genome resequencing of 15 maize inbreds, common variants (CV) and clustering analyses were applied to identify non-synonymous SNPs (nsSNPs) and corresponding candidate genes for drought tolerance.ResultsA total of 524 nsSNPs that were associated with 271 candidate genes involved in plant hormone regulation, carbohydrate and sugar metabolism, signaling molecules regulation, redox reaction and acclimation of photosynthesis to environment were detected by CV and cluster analyses. Most of the nsSNPs identified were clustered in bin 1.07 region that harbored six previously reported QTL with relatively high phenotypic variation explained for drought tolerance. Genes Ontology (GO) analysis of candidate genes revealed that there were 35 GO terms related to biotic stimulus and membrane-bounded organelle, showing significant differences between the candidate genes and the reference B73 background. Changes of expression level in these candidate genes for drought tolerance were detected using RNA sequencing for fertilized ovary, basal leaf meristem tissue and roots collected under drought stressed and well-watered conditions. The results indicated that 70% of candidate genes showed significantly expression changes under two water treatments and our strategies for mining candidate genes are feasible and relatively efficient.ConclusionsOur results successfully revealed candidate nsSNPs and associated genes for drought tolerance by comparative sequence analysis of 16 maize inbred lines. Both methods we applied were proved to be efficient for identifying candidate genes for complex traits through the next-generation sequencing technologies (NGS). These selected genes will not only facilitate understanding of genetic basis of drought stress response, but also accelerate genetic improvement through marker-assisted selection in maize.

Magnesium (Mg(2+)) is an essential macronutrient for plant growth and development, and the CorA/MRS2/MGT-type Mg(2+) transporters play important roles in maintaining Mg(2+) homeostasis in plants. Although the MRS2/MGT genes have been identified in two model plant species, Arabidopsis and rice, a comprehensive analysis of the MRS2/MGT gene family in other plants is lacking. In this work, 12 putative MRS2/MGT genes (ZmMGT1- ZmMGT12) were identified in maize and all of them were classified into five distinct subfamilies by phylogenetic analysis. A complementation assay in the Salmonella typhimurium MM281 strain showed that five representatives of the 12 members possess Mg(2+) transport abilities. Inhibition of ZmMGT protein activity using the hexaamminecobalt (III) (Co-Hex) inhibitor indicated that the ZmMGT protein mediated both low-affinity and high-affinity Mg(2+) transport in maize. A semi-quantitative reverse transcription-PCR (RT-PCR) analysis revealed that eight genes were constitutively expressed in all of the detected tissues, with one being specifically expressed in roots and three having no detectable expression signals. A quantitative RT-PCR analysis showed that some ZmMGT members displayed differential responses to Mg(2+) deficiency and aluminum (Al) stress. Furthermore, root growth inhibition and Mg(2+) accumulation analyses in two maize inbred lines, which conferred different levels of Al tolerance, revealed that ZmMGT proteins contributed to the Al resistance of the Al tolerance genotype. We hypothesize that ZmMGT family members function as Mg(2+) transporters and may play a role in linking Mg(2+) deficiency and Al stress responses. Our results will be valuable in a further analysis of the important biological functions of ZmMGT members in maize.

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Genetic relationship of a newly discovered teosinte from Nicaragua, Zea nicaraguensis with waterlogging tolerance, was determined based on randomly amplified polymorphic DNA (RAPD) markers and the internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA using 14 accessions from Zea species. RAPD analysis showed that a total of 5,303 fragments were produced by 136 random decamer primers, of which 84.86% bands were polymorphic. RAPD-based UPGMA analysis demonstrated that the genus Zea can be divided into section Luxuriantes including Zea diploperennis, Zea luxurians, Zea perennis and Zea nicaraguensis, and section Zea including Zea mays ssp. mexicana, Zea mays ssp. parviglumis, Zea mays ssp. huehuetenangensis and Zea mays ssp. mays. ITS sequence analysis showed the lengths of the entire ITS region of the 14 taxa in Zea varied from 597 to 605 bp. The average GC content was 67.8%. In addition to the insertion/deletions, 78 variable sites were recorded in the total ITS region with 47 in ITS1, 5 in 5.8S, and 26 in ITS2. Sequences of these taxa were analyzed with neighbor-joining (NJ) and maximum parsimony (MP) methods to construct the phylogenetic trees, selecting Tripsacum dactyloides L. as the outgroup. The phylogenetic relationships of Zea species inferred from the ITS sequences are highly concordant with the RAPD evidence that resolved two major subgenus clades. Both RAPD and ITS sequence analyses indicate that Zea nicaraguensis is more closely related to Zea luxurians than the other teosintes and cultivated maize, which should be regarded as a section Luxuriantes species.

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