QTL Mapping by Whole Genome Re-sequencing and Analysis of Candidate Genes for Nitrogen Use Efficiency in Rice.

Xinghai Yang,Faqian Xiong,Danting Li,Yanyan Wu,Guofu Deng,Ju Gao,Xiuzhong Xia,Baoxuan Nong,Yu Zeng,Zongqiong Zhang

doi:10.3389/fpls.2017.01634

Abstract

Nitrogen is a major nutritional element in rice production. However, excessive application of nitrogen fertilizer has caused severe environmental pollution. Therefore, development of rice varieties with improved nitrogen use efficiency (NUE) is urgent for sustainable agriculture. In this study, bulked segregant analysis (BSA) combined with whole genome re-sequencing (WGS) technology was applied to finely map quantitative trait loci (QTL) for NUE. A key QTL, designated as qNUE6 was identified on chromosome 6 and further validated by Insertion/Deletion (InDel) marker-based substitutional mapping in recombinants from F2 population (NIL-13B4 × GH998). Forty-four genes were identified in this 266.5-kb region. According to detection and annotation analysis of variation sites, 39 genes with large-effect single-nucleotide polymorphisms (SNPs) and large-effect InDels were selected as candidates and their expression levels were analyzed by qRT-PCR. Significant differences in the expression levels of LOC_Os06g15370 (peptide transporter PTR2) and LOC_Os06g15420 (asparagine synthetase) were observed between two parents (Y11 and GH998). Phylogenetic analysis in Arabidopsis thaliana identified two closely related homologs, AT1G68570 (AtNPF3.1) and AT5G65010 (ASN2), which share 72.3 and 87.5% amino acid similarity with LOC_Os06g15370 and LOC_Os06g15420, respectively. Taken together, our results suggested that qNUE6 is a possible candidate gene for NUE in rice. The fine mapping and candidate gene analysis of qNUE6 provide the basis of molecular breeding for genetic improvement of rice varieties with high NUE, and lay the foundation for further cloning and functional analysis.

Highlights

Nitrogen, one of the most demanding mineral elements in plants, is the most common limiting factor for plant growth in nature
The nitrogen use efficiency (NUE) was evaluated for each F2:3 families and for two lines (GH998 and NIL-13B4)
The NUE of the 280 F2 lines were fluctuated in the range of 3.64–34.39% with the maximum efficiency being 9.32 times of the minimum efficiency, the mean efficiency of 18.05, the standard deviation of 6.97, and the normality test value of 0.9821, indicating that the phenotype of NUE in F2 population is accorded with normal distribution (Figure 1)

Summary

Introduction

One of the most demanding mineral elements in plants, is the most common limiting factor for plant growth in nature. Hu et al (2015) showed that NRT1.1B had a nitrate-transporting activity at low and high nitrate ion concentrations. In rice, Lin et al (2000) cloned OsNRT1 gene and found that it was homologous to Arabidopsis thaliana AtNRT1 gene and encodes a low affinity nitrate transporter. Another nitrate transporter, NRT2, is a high-affinity nitrate transporter but can’t transfer NO−3 independently. Fan et al (2016) found that OsNRT2.3b was able to increase the pH-buffering capacity of the plant, increasing the uptake of N, Fe, and P, improving NUE and grain yield OsNAR2.1, a partner protein for the highaffinity nitrate transporter, is able to interact with OsNRT2.1, OsNRT2.2, and OsNRT2.3, and can enhance nitrate uptake by rice roots at different nitrate supply levels (Yan et al, 2011). Chen et al (2017) indicated that rice NO−3 uptake, yield and NUE were improved by increased OsNAR2.1 expression via its native promoter. Fan et al (2016) found that OsNRT2.3b was able to increase the pH-buffering capacity of the plant, increasing the uptake of N, Fe, and P, improving NUE and grain yield

Methods

Results

Discussion

Conclusion