Analysis of the genetic basis of plant height-related traits in response to ethylene by QTL mapping in maize (<i>Zea mays</i> L.)

<div><p>Ethylene (ET) is critical importance in the growth, development, and stress responses of plants. Plant hormonal stress responses have been extensively studied, however, the role of ET in plant growth, especially plant height (PH) remains unclear. Understanding the genetic control for PH in response to ET will provide insights into the regulation of maize development. To clarify the genetic basis of PH-related traits of maize in response to ET, we mapped QTLs for PH, ear height (EH), and internode length above the uppermost ear (ILAU) in two recombinant inbred line (RIL) populations of <i>Zea mays</i> after ET treatment and in an untreated control (CK) group. Sixty QTLs for the three traits were identified. Twenty-two QTLs were simultaneously detected under both ET treatment and untreated control, and five QTLs were detected at two geographic locations under ET treatment only. Individual QTL can be explained 3.87–17.71% of the phenotypic variance. One QTL (<i>q2PH9-1</i>, <i>q1PH9</i>, <i>q1EH9</i>/<i>q1ILAU9-1</i>, <i>q2ILAU9</i>, and <i>q2EH9</i>) for the measured traits (PH, EH, ILAU) was consistent across both populations. Two QTLs (<i>q2PH2-5</i>, <i>q2ILAU2-2</i>, <i>q1PH2-2</i>, and <i>q1ILAU2-2</i>; <i>q1PH8-1</i>, <i>q1EH8-1</i>, <i>q2PH8-1</i>) were identified for up to two traits in both locations and populations under both ET treatment and untreated control. These consistent and stable regions are important QTLs of potential hot spots for PH, ear height (EH), and internode length above the uppermost ear (ILAU) response to ET in maize; therefore, QTL fine-mapping and putative candidate genes validation should enable the cloning of PH, EH, and ILAU related genes to ET response. These results will be valuable for further fine-mapping and quantitative trait nucleotides (QTNs) determination, and elucidate the underlying molecular mechanisms of ET responses in maize.</p></div

Chromosomal locations of QTLs for plant height-related traits in Pop.

<p>2 (KUI3 × B77, N = 177 RILs) population across two environments and two ethylene treatments. Note: Rectangle QTL detected for plant height (PH) with ethylene treatment (Solid) or without (Hollow), Oval QTL detected for ear height (EH) with ethylene treatment (Solid) or without (Hollow), and Inverted triangle QTL detected for internode length above the uppermost ear (ILAU) with ethylene treatment (Solid) or without (Hollow). W indicates a QTL detected in Wuqiao test station, and L indicates a QTL detected in Lishu test station.</p

Combined analysis of variance for plant height-related traits in the ethylene treated and control RIL populations grown in two different locations (<i>F</i>-values).

<p>Combined analysis of variance for plant height-related traits in the ethylene treated and control RIL populations grown in two different locations (<i>F</i>-values).</p

Comparison of mapping results across the two RIL populations or ethylene treatments in two locations.

<p>Comparison of mapping results across the two RIL populations or ethylene treatments in two locations.</p

Correlation coefficients between plant height-related traits in the two RIL populations.

<p>Correlation coefficients between plant height-related traits in the two RIL populations.</p

Chromosomal locations of QTLs for plant height-related traits in Pop.

<p>1 (K22 × BY815, N = 197 RILs) population across two environments and two ethylene treatments. Note: Rectangle QTL detected for plant height (PH) with ethylene treatment (Solid) or without (Hollow), Oval QTL detected for ear height (EH) with ethylene treatment (Solid) or without (Hollow), and Inverted triangle QTL detected for internode length above the uppermost ear (ILAU) with ethylene treatment (Solid) or without (Hollow). W indicates a QTL detected in Wuqiao test station, and L indicates a QTL detected in Lishu test station.</p

Leaf water status assay.

<p>Changes in leaf water potential (A) and relative water content (B) in WT and transgenic maize (lines M-6 and M-8) subjected to drought stress. Drought stress treatment was imposed as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0052126#pone-0052126-g002" target="_blank">Fig. 2 (A)</a> above. Error bars denote the standard deviation values, and asterisks indicate a significant difference (*<i>P</i><0.05) compared with the corresponding controls.</p

Morphological characteristics of 21-d-old seedlings of WT and transgenic maize.

<p>The data point are the mean of two independent biological experiments, and each experiment comprised five samples. Error bars denote the standard deviation values.</p

Drought-stress-tolerant phenotype, water loss and stomatal assay.

<p>(A) Drought-stress-tolerant phenotype of transgenic maize (lines M-6 and M-8). Drought stress treatment was applied to 21-d-old seedlings of WT and transgenic maize by completely withholding irrigation for 10 d, then re-watering for 2 d. 21 d, 21-d-old seedling; D 2 d, completely withholding irrigation for 2 d; D 5 d, completely withholding irrigation for 5 d; D 10 d, completely withholding irrigation for 10 d; R 2 d, re-watered for 2 d. (B) Survival rates of transgenic maize plants overexpressing <i>LOS5</i> under drought-stress conditions. Fifty 21-d-old seedling from each lines or WT were deprived of water for 14 d, watering was resumed for 7 d, and then the plants were scored for viability. Plants were considered dead if all the leaves were brown and there was no growth after 7 d of watering. Each column represents an average of two independent experiments with three replicates, and values represented the mean ±SD. (C) Comparison of transpirational water loss in detached leaves of WT and transgenic maize. Values represent the mean ±SD (n = 4). (D) Comparison of stomatal apertures of WT and transgenic maize under drought stress conditions. Drought stress treatment was imposed as described in Fig. 2 (A) above. Error bars denote the standard deviation values, and asterisks indicated a significant difference (*<i>P</i><0.05) compared with the corresponding controls.</p

RT-qPCR analysis of stress-responsive genes.

<p>RNA levels of <i>Rad 17</i> (A), <i>NECD1</i> (B), <i>CAT1</i> (C), and <i>ZmP5CS1</i> (D) genes were determined by RT-qPCR using RNAs isolated from 21-d-old WT and transgenic maize (lines M-6 and M-8) exposed to 20% PEG. <i>Actin</i> gene was used as the control. The expression level of transgenic lines is shown relative to the expression of WT plants grown under well-watered condition. Error bars denote the standard deviation values, and asterisks indicate a significant difference (*<i>P</i><0.05; ** <i>P</i><0.01) compared with the corresponding controls.</p