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Nitrogen is one of the necessary nutrition for plant growth and development, and also one of the limited factors for yield. In the agricultural natural ecological system, the nitrogen mainly exist by three different forms:ammonium (NH4+-N), nitrate (N03-N) and ammonium nitrate (NH4+-NO3-). At present,70%of the world’s rice cultivation in paddy field and upland soil, due to long-term natural selection and evolution, lowland rice prefer ammonium as the major nitrogen source, and upland rice utilize nitrate for growth. In generally, high concentration of NH4+often inhibits seminal root elongation in upland cultivars. To elucidate mechanism underlying this inhibition by NH4+on root elongation, we carried out quantitative trait locus (QTL) and metabolome analyses using a mapping population derived from the cross between "Akihikari"(NH4+-tolerant lowland cultivar) and "IRAT109"(NH4+-susceptible upland cultivar), the main research results are as follows:1. Akihikari and IRAT109were hydroponically for9days in different concentration of three nitrogen forms, containing (NH4)2SO4、NH4NO3and Ca(N03)2, respectively. With increasing the concentrations of (NH4)2SO4and NH4NO3, Akihikari and IRAT109seminal root elongation inhibited, while Ca(N03)2have no effect on seminal root elongation, even in high concentration conditions.2. Akihikari and IRAT109were grown in500μKN03、(NH4)2SO4, and NH,N03as the sole nitrogen source treatment, all three kinds of nitrogen forms no significant effects on Akihikari seminal root elongation, however IRAT109with the longest seminal root in KNO3solution, followed by NH4NO3, and in (NH4)2SO4, has the shortest root. 3. Perform QTL analysis of genetic effects on lowland and upland rice seminal root elongation which treated in three different nitrogen forms (NO3--N、NH4+-N and NO3--NH4-),wo specific QTL regions were detected, Q7-1was located on chromosome7between the marker RM3859-RM5405, and IRAT109(upland rice) allele promote seminal root elongation in N03condition (upland soil),while Q2-1was located on chromosome2between marker RM3421-RM5472, and Akihikari (lowland rice)allele promote the seminal root elongation in NH4+condition (paddy field).4. When grown hydroponically over the wide ranges of NH4+concentration (20μM,200μM and2mM), Akihikari and IRAT109seedlings showed different growth responses to NH4+. In Akihikari seminal root length was almost uniform over the ranges of NH4-, while seminal root length was progressively reduced with increasing NH4+concentration in IRAT109. Such inhibitory effect on IRAT109roots was never seen in NO3-:seminal root elongation was uniform over the ranges of N03in both Akihikari and IRAT109. High concentration of NH4-was much inhibitorier for IRAT109root than for Akihikari root. In addition, through the accompaning ion test study shows that only NH4-inhibit seminal root elongation.5. Analysis BC1F11population with QTL Cartgrapher2.0revealed that a total of six genomic regions, distributed on chromosomes1,2,6(two QTLs),9and11, affected root elongation. The IRAT109allele promoted elongation on four (qSRL1-1, qSRL6-1, qSRL6-2and qSRL9-1) out of six QTLs, which consisted with the fact that IRAT109has a deeper root system than Akihikari. Comparison of QTLs between low and high NH4+treatments revealed that no QTLs were specific for low (20μM) NH4+treatment while two QTLs (qSRL2-1and qSRL6-2) were specific for high (2mM) NH4+treatment. Of the two, one (qSRL2-1) seemed to be involved in the growth inhibition by high NH4; only at this QTL the IRAT109allele reduced root growth under high concentration of NH4+ 6. Measurement of29amino acids content in Akihikari and IRAT109root were analyzed by capillary electrophoresis-mass spectrometry (CE-MS). At low concentration of NH4, amino acid composition was similar between "Akihikari" and "IRAT109", with glutamine as the major component followed by alanine, glutamate, serine and asparagine. Under high NH4+concentration, however, amino acid contents were fairly different between two cultivars; glutamine and glutamate were notably increased in "Akihikari" while serine was increased only in "IRAT109". Alanine and asparagine were increased similarly between the two cultivars.7. We determined QTLs controlling the content of these amino acids. Of these amino acids, serine seemed to be related to root elongation. We detected two QTLs for serine content on chromosomes2and11where the IRAT109allele increased serine content under low (chromosome11) and high (chromosome2) NH4-, respectively. Interestingly, these two QTL were co-located with the seminal root length QTLs with negative effect from IRAT109allele (qSRL2-1and qSRL11-1). Among them was the QTL on chromosome2(qSRL2-1) where the IRAT109allele reduced root growth under high concentration of NH4-, indicating that accumulation of serine under high concentration of NH4+might inhibit root elongation.Morover, additional of0.1mM or0.5mM of glutamine、glutamate、 asparagine、aspartate and serine to the growth medium to examined the effects of these amino acids on root growth. Addition of serine to growth medium severely inhibited seminal root elongation, while glutamine, glutamate, alanine, and asparagine had little effects on seminal root elongation. From these results, we concluded that inhibitory effect of serine may, at least in part, be involved in the inhibition of rice root elongation by NH4.
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