Table of Content

12 March 2010, Volume 36 Issue 3

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CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Integration and Insertion Site of EPSPs Gene on the Soybean Genome in Genetically Modified Glyphosate-Resistant Soybean

WANG Xiao-Bo,JIANG Ling-Xue,WEI Li,LIU Lin,LU Wei,Li Wen-Xin,WANG Jun,CHANG Ru-Z

Acta Agron Sin. 2010, 36(3):  365-375.  doi:10.3724/SP.J.1006.2010.00365

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The detection of genetically modified crops (GMCs) is becoming both food labels and legal necessity. The aim of this study was to identify integration and insertion locus of foreign EPSPs gene in genetically modified soybean for the safety assessment of genetically modified crop. The EPSPs gene fragment was detected using gene specific primes and immunochromatographic strip was used to detect the EPSPs protein. The result showed that the EPSPs gene was integrated into soybean genome and EPSPs protein could be expressed normally. Genome walking was used to analyze the flanking sequences of both 35S promoter and NOS terminator, and soybean genome database (Phytozome) was used to analyze the insertion site and study the effect of the insertion on soybean genome. The Dra I and EcoR V restriction enzyme were found that they could be used to digest the soybean genome completely, and the library with adaptor was established and nest PCR was used to amplify the flanking sequences of 35S and NOS genes in genetically modified soybean genome. The result showed that the start site of flanking region of either 35S promoter or NOS terminator was Gm02:7912740 or Gm02:7777705, respectively, which means the foreign gene may not insert into a fixed position, and one 135 kb DNA fragment may be translocated. Two unknown sequences and two soybean DNA fragments with physical distance of 24 kb in opposite directions were found at the flanking region of NOS terminator. We also found that there were high AT content (about 70%) and low gene density in 90kb flanking regions of the insertion locus, and one gene coding HEC1 and HEAT repeat domain (Glyma02g09790) was found to be rearranged. RT-PCR and qRT-PCR showed that the gene was down-regulated during PEG and ABA treatment. In this study, we found the genetically engineered soybean genome was rearranged because of the insertion of foreign genes and one gene coding HEC1 and HEAT repeat domain (Glyma02g09790) which may response to drought stress through ABA signal pathway was identified for the first time.

Comparison between QTLs for Chlorophyll Content and Genes Controlling Chlorophyll Biosynthesis and Degradation in Japonica Rice (Oryza sativa L.)

JIANG Shu-Kun,ZHANG Xi-Juan,XU Zheng-Jin,CHEN Wen-Fu

Acta Agron Sin. 2010, 36(3):  376-384.  doi:10.3724/SP.J.1006.2010.00376

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To understand the expression patterns of chlorophyll content related genes at different stages and genetic mechanisms of stay-green at later stage, we analyzed the QTLs controlling chlorophyll content at the stages of tillering, heading and maturity by employing 126 recombinant inbred lines (RILs) derived from a cross between two japonica rice cultivars, Shennong 265 and Lijiangxintuanheigu. Five, seven and ten QTLs controlling chlorophyll contents at tillering stage, heading stage and maturity stage were detected, respectively. They were distributed on all rice chromosomes except chromosome 5. Comparison of the QTLs and the genes underlying the key enzymes of chlorophyll biosynthesis and degradation revealed that relatively more QTLs detected at earlier stage co-located with the genes related to chlorophyll biosynthesis and degradation. With the growth stage going on, more QTLs were detected but only a few of them involved in chlorophyll biosynthesis and degradation. The results suggested that the expression level of most genes related to chlorophyll biosynthesis (degradation) had no difference at earlier stage but specific key genes increased at later stage. And two possible genetic bases for stay-green were proposed.

Genetic Improvement of Wheat Growth Habit and Its Molecular Marker-Assisted Selection in Yellow and Huai River Reaches

ZHANG Jing,WU Suo-Wei,LIU Bing-Hua,SONG Mei-Fang,ZHOU Peng,GUO Chun-Yan,et al.

Acta Agron Sin. 2010, 36(3):  385-390.  doi:10.3724/SP.J.1006.2010.00385

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The vernalizatioin gene in wheat cultivar Shimai 12 (Triticum aestivum L.) was investigated to find a more efficient way for the improvement of growth habit of wheat cultivars grown in Yellow and Huai River Reaches. In the F_2:3 lines derived from the cross between Shimai 12 and Shijiazhuang 8 (a winter wheat cultivar), the ratio of the spring, segregating, and winter lines was in accordance to 1:2:1 based on chi-square test, showing that the spring growth habit of Shimai 12 was determined by single dominant gene, which was identified as Vrn-D1 by gene-specific markers. The Vrn-D1 gene-specific marker was further used to genotype the F_2:3 lines, and the results were consistent with phenotyping, verifying the available utilization of the marker assisted selection for Vrn-D1 in breeding programs. Besides, summer sowing in high altitude and cool area proved to be a cost-saving and efficient method for wheat growth habit identification.

Genetic Diversity and Relationship of Tea Germplasm in Yunnan Revealed by ISSR Analysis

LIU Ben-Ying,LI You-Yong,TANG Yi-Chun,WANG Li-Yuan,CHENG Hao,WANG Ping-Sheng

Acta Agron Sin. 2010, 36(3):  391-400.  doi:10.3724/SP.J.1006.2010.00391

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Tea is an important beverage crop in the world. Yunnan region in China is the origin center of tea plants. Our knowledge on genetic diversity and relationship of tea germplasm in Yunnan province is critical to guide tea breeding. In order to provide theoretical information for using tea germplasm in tea breeding, we investigated the relationship and genetic diversity of the tea germplasm in Yunnan province. Eight species, including 134 tea varieties were used to detect the genetic variation by inter-simple sequence repeats (ISSR) analysis. The result showed that 475 DNA fragments among all 134 tea accessions were amplified, using 18 reliable ISSR primers, among which 470 DNA bands were polymorphic (PPB=98.9%). This indicated that a great amount of genetic polymorphism exists among tea germplasm tested. The genetic similarity (GS) among the tested genotypes ranged from 0.445 to 0.819, with an average of 0.512, indicating a wide gene pool among tea varieties in Yunnan. The cluster analysis presented that these resources were divided into three main groups using the unweighted pair-group method with arithmetic average (UPGMA) based on ISSR molecular marker data, but the dendrogram did not indicate clear division among tested varieties based on their geographical origin. Principal Component analysis (PCA) for ISSR data showed that PCA supported UPGMA clustering result, but showed more explicit relationships among the test accessions with different lays, orientationsand positions. The GS among 8 populations ranged from 0.850 to 0.987, with an average of 0.92, indicating that there existed a small variation of genetic diversity among different population. The findings of this research would be favorable for the further practice, such as tea breeding, the molecular genetic linkage mapping and the DNA fingerprint building of tea germplasm.

cDNA-AFLP Analysis Reveals Differential Gene Expression in Wheat Adult-Plant Resistance to Stripe Rust

ZHANG Gang,DONG Yan-Ling,XIA Ning,ZHANG Yi,WANG Xiao-Jie,QU Zhi-Peng,LI Yi-Min,

Acta Agron Sin. 2010, 36(3):  401-409.  doi:10.3724/SP.J.1006.2010.00401

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Wheat (Triticum aestivum L.) stripe rust is one of the most devastating diseases of wheat throughout the world. Adult plant resistance (APR) to stripe rust in wheat conferring durable resistance, thus, plays a pivotal role in the control of the disease. In the present study, to elucidate molecular mechanism of wheat APR to stripe rust, we conducted extensive transcription profiling of adult-plant wheat cultivar Xingzi 9104 infected by Puccinia striiformis Westend f. sp. tritici Erikss. pathotype CY32 using cDNA-AFLP technique. We analyzed transcription profiling of the incompatible reaction across nine sampling time points within five days after inoculation. Of the total 32,320 transcript derived fragments (TDFs) obtained using cDNA-AFLP with 64 primer pairs, 2201 (6.81%) displayed altered expression patterns after inoculation, of which 926 showed up-regulated and 1275 down-regulated. Three hundred and thirty differentially expressed TDFs produced reliable sequences after cloning and sequencing, of which 259 expressed sequence tags (ESTs) of unigenes were obtained after assembling, designated from aTaPST1 to aTaPST259, deposited in GenBank with accessions numbers from FL645754 to FL646011 and FL646262. BLASTX analyses and functional annotations were then performed and the results revealed that the 95 ESTs had predicted gene products mainly implicated in energy (11.20%), metabolism (4.63%), transcription (3.86%), disease/defense (3.86%), protein destination and storage (3.09%), protein synthesis and cell growth (each accounted for 2.32%), and signal transduction (1.54% of the sequenced total 259 ESTs). Six differential genes related to disease/defense, transcription, and signal transduction were chosen for further qRT-PCR expression patterns, which confirmed the cDNA-AFLP profiles. Our results indicated that wheat APR to stripe rust involved in multifaceted biochemical and physiological reactions, including concerted regulation of the genes involved in different pathways like disease/defense, transcription, protein metabolism, signal transduction, as well as abiotic stresses. These results provide information for further elucidation of molecular mechanism of wheat APR to stripe rust.

Development and Utiligaiton of Orthologous SSR Markers in Arachis through Soybean (Glycine max) EST

HONG Yan-Bin,CHEN Xiao-Ping,LIU Hai-Yan,ZHOU Gui-Yuan,LI Shao-Xiong,WEN Shi-Jie

Acta Agron Sin. 2010, 36(3):  410-421.  doi:10.3724/SP.J.1006.2010.00410

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Lack of sufficient molecular markers hindered current genetic research in peanuts (Arachis hypogaea L.). It is necessary to enrich molecular markers for potential use in peanut breeding programs. Recently, EST-SSRs have received much attention with the higher level of transferability to closely related species and it can be rapidly developed from EST database by data mining at low cost. In this study, we mined SSRs from the soybean EST and analyzed their transferability in peanuts (Arachis hypogaea L.) and its wild species. A total of 394 370 soybean ESTs were clustered to generate 82 614 Uni-ESTs, 2 082 ESTs of which contained 2 191 SSRs with 1 SSR in 22.96 kb EST. Di-nucleotide motif (63.5%), followed by tri-nucleotide (30.9%), were the most abundant in soybean EST-SSR. The top two motif sequence types with high frequency were AG/CT (35.8%) and AT/AT (25.4%). Based on the 2082 SSR-ESTs, a total of 685 primer pairs were successfully designed and synthesized to test the amplification in four soybean cultivars. There were 582 primer pairs amplified effectively in soybean cultivars, 98 of which exhibited polymorphism. The cross-transferability of soybean EST-SSR was different among nine sections in Arachis, ranging from 12.4% to 15.7% with an average of 14.2%. Polymorphism analysis in Arachis section showed that 69 of the 79 orthologous SSR markers displayed polymorphsim in ten wild species, while only five markers had polymorphism in 22 cultivars. The PCR bands amplified in soybean and Arachis section by primer ES-105 were cloned and sequenced. The results revealed that the polymorphism among two soybean cultivarswas caused by little variation in SSR motif (AT)n, while the polymorphsim between soybean and Arachis section was contributed not only by the indel in SSR loci, but also by the frequence of the indel and substitution in franking region.The work indicated that the development of orthologous SSR markers for Arachis from soybean EST is feasible. There was a high level of polymorphism in Arachis sections for soybean-derived EST-SSR, therefore, the research of comparative genomics of Glycine and Arachis can be performed due to the high transferability of functional molecular markers.

Genetic Mapping of a Bacterial Blight Resistance Gene Xa14 in Rice

BAO Si-Yuan,TAN Ming-Pu,LIN Xing-Hua

Acta Agron Sin. 2010, 36(3):  422-427.  doi:10.3724/SP.J.1006.2010.00422

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Bacterial blight of rice, which is a vascular bundle disease caused by Xanthomonas oryzae pv. oryzae (Xoo), is one of the most serious diseases of rice worldwide, and leads to the yield of rice reduced greatly. The dominant gene Xa14 is highly resistant to Philippine race 5 of bacterial blight. The gene was located at the distal end on the long arm of chromosome 4 by Taura et al. Near-isogenic line in the background of IR24 namely IRBB14 carrying the geneXa14has been developed at International Rice Research Institute(IRRI). In order to construct the high-resolution likage maps for the Xa14 region on chromosome 4 for finally clone by positional cloning, tow F₂ populations were used to estimate linkage based on marker genotype and reaction to disease inoculation with Xanthomonas oryzae pv. oryzae. Using 145 highly susceptible individuals from a total of 775 plants of F₂ population of the cross between IRBB14 and IR24, the gene Xa14 was located in a 0.68 cM region on the nearby end of chromosome 4, which was flanked by the molecular markers, HZR970-8 and HZR988-1, with the distance of 0.34 cM between the flanked markers and Xa14, respectively, and completely cosegregated with the SSR markers, HZR645-4, HZR669-2, HZR669-5, and HZR669-7 in this population. Using 158 highly susceptible individuals from a total of 763 plants of F₂ population of the cross between IRBB14 and ZZA, the gene Xa14 locus was mapped to the interval between HZR648-5 and RM280, and was 1.90 cM away from HZR648-5 which was the closest marker flanking the Xa14 locus. Combining recombination frequencies for the two populations together, the gene Xa14 was mapped to 3 BAC clones spanned approximately 300 kb in length between SSR markers HZR970-8 and HZR988-1.

Development and Evaluation of Peanut Germplasm with Resistance to Aspergillus flavus from Core Collection

JIANG Hui-Fang,REN Xiao-Ping,WANG Sheng-Yu,ZHANG Xiao-Jie,HUANG Jia-Quan

Acta Agron Sin. 2010, 36(3):  428-434.  doi:10.3724/SP.J.1006.2010.00428

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Peanut (Arachis hypogaea L.), one of the main oil and cash crops in the world, is easily susceptible to Aspergillus flavus, resulting huge loss in its quality, so Aspergillus flavus infection greatly limits peanut production and industry in China. Therefore, it is imperative to develop new peanut germplasm with resistance to Aspergillus flavus in breeding program. The core collection is well accepted as a useful way to improve the efficiency of crop germplasm evaluation and utilization, which contains a subset of accessions from the entire collection that covers the most of available genetic information. In the present study, a total of 561 accessions of Chinese peanut core collection and 155 accessions of ICRISAT mini core collection were identified. Eight varieties with resistance to Aspergillus flavus invasion and aflatoxin production each were developed, including one (51002-6) with elite agronomic traits. The peanut germplasm with resistance to Aspergillus flavus invasion and aflatoxin production in ICRISAT mini core were more than those in Chinese peanut core collection. In addition, the percentages of accessions with resistance to Aspergillus flavus invasion in var. hypogaea, and accessions resistant to aflatoxin production in var. hirsuta were relatively high in comparison with others. Genetic diversity in the resistant peanut selections was evaluated based on morphological traits and SSR approach. ICG12625 with resistance to aflatoxin production and ICG4750 with resistance to aflatoxin invasion were evaluated by SSR, the genetic distance of them with high-yielding cultivars such as Zhonghua 5, Zhonghua 6 and Zhonghua 12 and Yuanza 9102 was larger. The primers were designed based on the conserved NBS-LRR domains of the disease resistance genes sequence, one RGA (Resistance gene analog) from genomic DNA of six different peanuts with resistance to Aspergillus flavus was obtained through PCR.

Cloning and Characterization of Six Oleic Acid Desaturase Pseudogenes of Brassica napus

XIAO Gang,ZHANG Zhen-Qian,WU Xian-Meng,TAN Tai-Long,GUAN Chun-Yun

Acta Agron Sin. 2010, 36(3):  435-441.  doi:10.3724/SP.J.1006.2010.00435

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The phenomenon of multi-copy genes is common in plants. Pseudogene is defined as an inactive gene, which can not synthesize functional proteins but share the similar DNA sequences with normal functional genes. In this study, 56 FAD2 DNA clones and 47 FAD2 seed cDNA clones of Brassica napus cv. Xiangyou 15 were investigated, and 6 new copies of FAD2 were detected, designated as FAD2P1-6 respectively. This sequence length of 6 copies ranged from 1 141–1 157 bp and there were no introns in their open reading frames (ORF). These 6 copies share 96.1% identity in nucleotides from one another, and share more than 87% nucleotides identity with AY577313. Deduced amino acid sequences revealed that 1–12 stop codons occurred in the coding region of six copies which will prevent them from coding for a functional protein. These six copies were investigated in vivo in Saccharomyces cerevisiae through being cloned into yeast expression vector pYES2.0, and the 16:2 and 18:2 fatty acids were determined by gas chromatographic analysis. The results revealed that the products of the six copies were not able to synthesize 16:2 and 18:2 fatty acids, suggesting that they are pseudogenes of FAD2. These multiple pseudogenes of FAD2 within the B. napus genome might result from the duplication of large chromosomal segments simultaneously following mutation. Because of the existence of multiple pseudogenes for FAD2 in B. napus genome, we should be careful in genetic research to identify true and false, to avoid wrong conclusions.

Analsysis of QTLs for Root Traits at Seedling Stage Using an "Immortalized F₂"Population in Wheat

LI Zhuo-Kun,PENG Tao,ZHANG Wei-Dong,XIE Quan-Gang,TIAN Ji-Chun

Acta Agron Sin. 2010, 36(3):  442-448.  doi:10.3724/SP.J.1006.2010.00442

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Identification of quantitative trait loci (QTLs) of roots plays an important role in the genetic mechanism study of root system. A number of QTLs for root traits have been detected using different populations in rice (Oryza sativa L.), but rarely in wheat (Triticum aestivum L.). The “immortalized F₂” population is favorable for QTL analysis due to their similar proportions of genotypes in the population, and the replicability at multiple locations over several years. The objective of this study was to detect QTLs for several root traits at seedling stage in wheat. From a set of doubled haploid lines derived from Huapei 3 × Yumai 57, an “immortalized F₂” population was constructed with 168 single crosses. Wheat seedlings were hydroponically cultured with Hoagland’s solution in a light incubator, and sampled when the fifth leaf emerged. Nine root traits including root total length, root surface area, root average diameter, root volume, root tips, maximum root length, shoot dry weight, root dry weight, and ratio of root dry weight to shoot dry weight per plant were measured using the WinRHIZO Root Analysis System. QTLs associated with the nine traits were detected using composite interval mapping (CIM) method. A total of seven additive QTLs and twelve pairs of epistatic QTLs were mapped on chromosomes 1A, 1D, 2A, 2B, 2D, 3A, 3B, 5D, 6D, and 7D. Additive, dominant and epistatic effects were observed across these QTLs, including the interactions between additive and additive, additive and dominance, dominance and additive, as well as dominance and dominance. The phenotypic variation explained by each QTL ranged from 0.01% to 11.91%. In the interval between XWMC41 and XBARC349.2 on chromosome 2D, a QTL for total root length and the root dry weight was detected. Epistasis was of great importance in root growth and development at seedling stage in wheat. Theresults showed a complex mechanism on the genetics of root traits in wheat, and the eight root traits were significantly correlated with each other. In marker-assisted breeding practice, several root traits should be considered simultaneously for an integrated performance of roots with proper size and shape.

TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY

Variation in Photosynthetic Traits and Antioxidant Enzyme Activities of Wheat Seedlings Transferred from Low to High Light Growth Condition

LI Hong-Wei,LI Bin,ZHENG Qi,LI Zhen-Sheng

Acta Agron Sin. 2010, 36(3):  449-456.  doi:10.3724/SP.J.1006.2010.00449

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When plants absorb excessive light energy, a large number of reactive oxygen species is generally produced resulting in the degradation of DNA, proteins, and pigments in plants. For wheat (Triticum aestivum L.) grown in North China, the photooxidation induced by high light (HL) during grain-filling period usually causes great losses in grain yield. It is important to understand the mechanism of wheat plant in response to HL for HL-tolerant breeding in wheat. Xiaoyan 54, a wheat cultivar with high resistance to HL, is an ideal material to disclose photosynthesis characteristics of wheat when exposed to HL. In this study, the third-leaf seedlings of Xiaoyan 54 were grown under the condition low light to HL in a growth chamber. The seedlings were sampled at 0, 1, 3, 8, 24, and 48 h of HL treatment. Simultaneously, the net photosynthetic rate (P_n), chlorophyll content (Chl), and fluorescence parameters were measured with the second leaf. The activities of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR), were also determined. In addition, expression pattern of the pigment binding protein related genes were evaluated. The results showed that P_n increased in the photosynthetic induction stage that was from 0 h to 8 h of HL treatment, but decreased continuously during the photoinhibition stage that was from 8 h to 48 h of HL treatment. The maximum P_n value of 18 μmol CO₂ m^-2 s^-1 was observed at the 8 h timepoint of HL treatment. The parameters G_s, C_i, and T_r changed similarly to P_n and reached the peaks at the 8 h timepoint of treatment. The contents of total chlorophyll and chlorophyll a only changed slightly during the 48 h of HL treatment. In contrast, chlorophyll b reduced significantly from 24 h to 48 h of treatment, and the ratio of chlorophyll a/b increased from 8 h to 48 h of HL treatment. At one hour of HL treatment, the maximum quantum efficiency of PSII (F_v/F_m), the maximum fluorescence (F_m), and variable fluorescence (F_v) were down-regulated significantly, when the heat dissipation was enhanced. The activities of SOD, CAT, APX, and GR were induced to higher levels with the highest value at 24 h timepoint of HL treatment. From 8 h to 48 h of HL treatment, the Talhcb genes, encoding LHCII subunits, were down-regulated at the RNA levels. At the early stage of HL treatment (0–3 h), TaELIP1 and TaELIP3 were induced, but repressed from 8 h to 48 h. As key enzymes in xanthophyll cycle, the transcripts of TaVDE and TaZEP responded differently to HL treatment. The expression of TaVDE decreased remarkably at 8 h of HL treatment and maintained a rather low level till 48 h. However, the expression of TaZEP showed an increase trend from 3 h to 24 h, and decreased at 48 h. In conclusion, when wheat seedlings exposed to continuous HL for 48 h, photooxidative stress occurred resulting in reductions of P_n, F_v/F_m, Chl b, and the expressions of pigment binding protein genes, but activation of the antioxidantenzymes.

Effects of Supplemental Irrigation Based on Testing Soil Moisture on Dry Matter Accumulation and distribution and Water Use Efficiency in Winter Wheat

HAN Zhan-Jiang,YU Zhen-Wen,WANG Dong,ZHANG Yong-Li

Acta Agron Sin. 2010, 36(3):  457-465.  doi:10.3724/SP.J.1006.2010.00457

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Water shortage is a serious problem threatening sustainable agricultural development in the North China Plain (NCP), where winter wheat (Triticum aestivum L.) is the largest water-consuming crop. Water-saving technique is one of the most important components in wheat cultivation system in this area. The purpose of this study was to optimize irrigation scheduling for high yield and water use efficiency (WUE) in wheat. Unlike earlier studies in which fixed irrigation amounts were given, we designed a ladder of relative soil moisture content at critical growth stages of wheat. The results are expected to provide general and valuable guidelines to farmers and irrigation managers in high-yielding wheat production in the NCP. The field experiments was conducted with the cultivar of Jimai 22 in Shiwang village (35.41°N, 116.41°E), Yanzhou, Shandong, China in 2007–2008 and 2008–2009 growing seasons. The irrigation treatments were designed based on the contents of relative soil moisture at sowing, jointing, and anthesis stages which were 80%, 65%, and 65% for treatment W0; 80%, 70%, and 70% for treatment W1; 80%, 80%, and 80% for treatment W2; 90%, 80%, and 80% for treatment W3, respectively. The accumulation and distribution of dry matter and water use efficiency (WUE) in wheat plants were investigated subject to soil moisture and supplemental irrigation. The results showed that dry matter accumulation amount in treatment W0 was the lowest whereas that in treatment W1 was the highest at maturity stage. The grain dry matter ratio was significantly higher in treatment W1 than in treatments W2 and W3. After anthesis, the redistribution amount and the ratio of dry matter that stored in vegetative organs before anthesis were presented as W0>W3>W2>W1, and the contribution of dry matter accumulation amount after anthesis to grains as W1>W2>W3>W0. Under the W1 condition, the filling rate and net photosynthetic rate maintained a relative high level at the end of filling stage, which was favorable for increasing the accumulation and distribution ratio of dry matter and the grain weight at maturity. The WUE in treatment W0 was higher than that in other treatments. However, the grain yield was the lowest in treatment W0. In both growing seasons, the grain yield, irrigation water use efficiency (WUE_I), precipitation use efficiency (WUE_P), and irrigation benefit (IB) in the three irrigation treatments decreased significantly as more water was supplied. Under the experimental condition, the W1 regime was considered as the optimal irrigation treatment, whose relative soil moisture content80% at sowing, 70% at jointing, and 70% at anthesis stage. When 43.8 and 13.8 mm of water was supplied in the 2007–2008 and 2008–2009 growing seasons, the final grain yield reached the highest level of 8 837.8 kg ha^-1 for 2007–2008 and 9 040.9 kg ha^-1 for 2008–2009 with the highest WUE_I and WUE_P.s in the 0–140 cm soil layer were

Effects of Different Cultivation Patterns on the Yield and Physiological Characteristics in Mid-Season Japonica Rrice

XUE Ya-Guang,CHEN Ting-Ting,YANG Cheng,WANG Zhi-Qin,LIU Li-Jun,YANG Jian-Chang

Acta Agron Sin. 2010, 36(3):  466-476.  doi:10.3724/SP.J.1006.2010.00466

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Rice is one of the most important food crops in China. The realization of continuously high yield has great significance in ensuring food security and society stability. Meanwhile, low nitrogen (N) use efficiency is a serious problem in rice production in China. The objective of this study was to investigate if a cultivation technique could coordinately increase both grain yield and N use efficiency in rice. Mid-season japonica rice cultivars were used with the treatments of local high-yielding cultivation (control), super high-yielding cultivation, and high-yielding and high N use efficiency cultivation (HHC). The characteristics of grain yield formation under different cultivation systems and their physiological basis were analyzed. The results showed that, compared with the control, the HHC significantly increased biomass of root and shoot, root cytokinin concentration and root oxidation activity, grain-leaf ratio, leaf photosynthetic rate during the mid and late grain filling periods, nonstructural carbohydrate accumulation in stems and sheaths at the heading time, remobilization of carbon accumulated from heading to mature stage, and harvest index. The HHC increased grain yield by 31% and agronomic N use efficiency (increased grain yield per unit N application) by 57% when compared with the control. The results suggest that root and shoot growth could be improved and high grain yield and high N use efficiency could be achieved through integrating and optimizing cultivation techniques in rice production.

Root Exudates and Nitrogen Uptake of Wheat in wheat/Maize/Soybean relay Cropping system

YONG Tai-Wen,CHEN Xiao-Rong,YANG Wen-Yu*,XIANG Da-Bing,FAN Gao-Qiong

Acta Agron Sin. 2010, 36(3):  477-485.  doi:10.3724/SP.J.1006.2010.00477

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Root exudates play a role in adjusting physical and chemical characteristics of soil, improving soil fertility, and facilitating nutrient uptake. Thus, they are used to explaining the mechanism of high nutrient utilization efficiency for plants. In the relay cropping system of wheat/maize/soybean, nitrogen use efficiency appears higher than that in the cropping system of wheat/maize/sweet potato. For explaining the relationship between the high use efficiency of nitrogen and the effects of rhizosphere microenvironment in the wheat/maize/soybean cropping system, the characteristics of wheat root exudates as well as its effects on wheat roots, wheat rhizosphere soil, and nitrogen uptake in wheat were studied in a two-year continuous experiment from 2006 to 2008 with four cropping systems, including two sole cropping systems (wheat–soybean and wheat–sweet potato) and two rely cropping systems (wheat/maize/soybean and wheat/maize/sweet potato). In the wheat/maize/soybean system, the soil moisture, soil pH and soil quick-acting N (NO₃⁻-N and NH₄⁺-N) content at wheat rhizosphere reduced at both flowering and maturity stages, and the total nitrogen uptake in shoots, root activity, root dry matter weight and soil total nitrogen content of wheat increased. At flowering stage, the total amount of organic acids and the content of soluble sugar of wheat rhizosphere increased in the wheat/maize/soybean system. The total amount of organic acid and the content of soluble sugar of wheat were higher in relay cropping system than in sole cropping system. In the two sole cropping systems, the wheat–soybean mode had higher amount of organic acid and higher content of soluble sugar than wheat–sweet potato. Wheat plants from fringe rows could exude organic acids and higher soluble sugar content than those from central rows, and the amount of root exudates in fringe rows was the highest among all the cropping systems. At jointing stage, the content of acetic acid in the sole cropping systems increased, which accounted for 47.8% to 51.6% of the total of organic acid. The content of citric acid in the relay cropping systems increased from 31.7% to 55.1% of total organic acid. At flowering stage, the content of acetic acid in sole and relay cropping systems was also promoted from 33.3% to 78.3% of the total organic acid. The root exudates were in favor of improving root growth condition, activating soil nutrition, enhancing nitrogen uptake and increasing nitrogen use efficiency in the wheat/maize/soybean relay cropping system.

CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Protein Accumulation in Grains of Wheat Cultivars Differing in Drought Tolerance and Its Regulation by Nitrogen Application Amount under Irrigated and Dryland Conditions

SUN Min,GUO Ping-Yi*,GAO Zhi-Qiang,WANG Peng,SHI Jing,MIAO Guo-Yuan

Acta Agron Sin. 2010, 36(3):  486-495.  doi:10.3724/SP.J.1006.2010.00486

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The protein accumulation in wheat (Triticum aestivum L.) grains is seldom studied under dryland condition although it has been widely tested under irrigated conditions. For understanding the protein accumulation characteristics of wheat cultivars differing in drought tolerance and the effects of water and nitrogen, a drought-tolerant cultivar, Jinmai 47, and a drought-sensitive cultivar, Nongda 189, were planted under both irrigated and dryland conditions with three nitrogen application rates. From the 5th day after anthesis, spikelets and flag leaves were sampled at a 5 days interval for measuring the contents of protein and its components in grains and the enzymes activities in grains and flag leaves. The results indicated that the contents of albumin, gliadin, glutenin, total protein, and the ratio of glutenin to gliadin in dryland cropping were higher than those in irrigated cropping, but the globulin content was lower than that in irrigated cropping. For Nongda 189, the contents of protein and its components and the ratio of glutenin to gliadin in grains were affected significently by drought, whereas for Jinmai 47, the variations on contents of globulin, gliadin, total protein, and ratio of glutenin to gliadin were slightly. The activities of glutamine synthetase (GS), glutamate synthase (GOGAT), glutamic acid-pyruvate transaminase (GPT) in grains and the activities of GS and GOGAT in flag leaves were reduced in dryland cropping compared with those in irrigated cropping. Simultaneously, the trend of GPT activity in grains was also affected by dryland cropping. The effect of drought on activities of these enzymes that involved in protein accumulation was greater in Nongda 189 than in Jinmai 47. Nitrogen application promoted the contents of protein and its components in grains, and higher rate of nitrogen application resulted in greater positive effect. Jinmai 47 presented greater effect of nitrogen application on protein content in grains than Nongda 189. Under both growing conditions, the activities of GS, GOGAT, and GPT in grains and the activity of GOGAT in flag leaves in all nitrogen treatments significantly correlated with the protein yield rather than the protein content in grains. The correlation between GS activity in flag leaves and protein yield in grains was different in the two cultivars. The above results suggest that the drought-tolerant cultivar is less affected by water condition compared with the drought-sensitive cultivar; application of nitrogen could promote the protein content in grains, and the effect is greater in the drought-tolerant cultivar than in the drought-sensitive cultivar.

TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY

Seed Growth Characteristics of Some Short Season Indeterminate Soybeans

Stephen J HERBERT, LIU Xiao-Bing,Gurkirat BAATH,JIN Jian,ZHANG Qiu-Ying,et al

Acta Agron Sin. 2010, 36(3):  496-501.  doi:10.3724/SP.J.1006.2010.00496

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The potential yield of a soybean crop is very specific to the genotype and the environment in which it is grown. For a given genotype, the interaction of density with arrangements of plants will play a key role in determining the competition for available resources especially solar radiation, water and nutrients, and thus maximum yield. Inter-plant competition begins when the immediate supply of a single necessary factor falls below the combined demands of all plants. Variation in yield among fields or years will be related to such inter-plant competition and borne out in one or more of the seed yield components of, plant per unit area, pods per plant, seeds per pod or weight per seed (seed size). This paper discusses the yield component of seed size in regulating differences in seed yield with changes in density and row spacing, and differences in seed yield between different years. Several field studies are reviewed where seed size was shown to be similar between nodes on a plant and in pods with varying seed number. However, the basal seed in two and three-seeded pods for many varieties tested was approximately 10% smaller than the middle or terminal seeds. Cotyledon cell number in basal seeds was 10% less than in middle or terminal seeds and all had similar cotyledon cell size. To test this further, we changed seed size through artificially source-sink manipulation, and by light enrichment and shading. Differences in seed size were also related to differences in cotyledon cell number. This relationship of seed size to cell number is intriguing since with induced changes in seed size, seeds at all nodes were of similar size even though lower nodes began filling 15 to 20 days before upper nodes.

Image Segmentation Technique of Field Cotton Based on Color Threshold

WANG Ling,WANG Ping,CHEN Bing-Lin,LIU Shan-Jun,JI Chang-Ying

Acta Agron Sin. 2010, 36(3):  502-507.  doi:10.3724/SP.J.1006.2010.00502

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The goal of cotton production in China is to improve corresponding rate of cotton quality grade; foreign fibers, adulteration, and cotton baling inconsistent phenomenon to decrease continuously. With the background, machine vision and pattern recognition technologies are introduced into traditional picking task to discriminate maturity degree and grade of quality of field cotton, which will solve the problem of picking cotton by the way from source, so that various cotton varieties can be adapted, pollution caused by agriculture chemicals can be avoided, labor cost can be reduced and agriculture cost can be decreased. In order to segment field cotton images exactly, we regarded cotton and its background as two classes and segmented them based on their color threshold. A total of 20 000 white, yellow, and stain cotton pixels and 20 000 background pixels of soil and cotton plant, including cotton bracteole, leaf, and branch, were extracted from typical under-ripe cotton images and ripe/over-ripe cotton images with various quality grades from 1 to 7. Color threshold of two classes of cotton and its background pixels were obtained in RGB, HSI, La*b*, and Hunter color space respectively; on the basis of which cotton regions were segmented from images; and HSI and La*b* color spaces were selected respectively by using S below 28, I over 108, L over 118, a* from 123 to 134, b* below 136 with less segmentation noise which would be removed based on morphological filter. The experiment results showed that 907 cotton images were segmented with an accuracy of 87.21% and 86.33% in HSI and La*b* color space respectively. The front images were segmented with an accuracy of 90.83% and 89.98% and the side images with an accuracy of 83.33% and 82.42%. Ripe cotton images were segmented perfectly in HSI color space while under-ripe cotton images in La*b* color space, and the speed-based segmentation method with threshold covering a wide area was preferable for field cotton surroundings.

Interaction Effects of Light Intensity and Nitrogen Supply on Gas Exchange, Some Enzyme Activities in Carbon-Nitrogen Metabolism and Quality in Flue-Cured Tobacco

YUN Fei, LIU Guo-Shun*,SHI Hong-Zhi

Acta Agron Sin. 2010, 36(3):  508-516.  doi:10.3724/SP.J.1006.2010.00508

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As a main nutritive element of tobacco, nitrogen not only plays an important role in protection against light stress, but also impacts on the balance of carbon-nitrogen metabolism. The combination of proper light intensity and nitrogen application could improve the utilization of light energy and increase the amount of dry matter accumulation in flue-cured tobacco. The objectives of this study were to reveal the interaction effects of light intensity and nitrogen application on gas exchange parameters, some enzyme activities in carbon-nitrogen metabolism and chemical composition and to investigate the optimum combination of light and nitrogen in tobacco growth. Flue-cured tobacco Yuyan 5 was used in a pot experiment in 2008, with the treatments of light intensity (four levels) and nitrogen supply (three treatments). The results indicated that the net photosynthetic rate (P_n), transpiration rate (T_r), stomatal conductance (G_s) and the amount of dry matter accumulation reduced under shading, but the intercellular CO₂ concentration (C_i) increased. Light, nitrogen and their interactions had a significant influence on photosynthetic physiology indicators of flue-cured tobacco. The decline in photosynthetic rate caused by shading could be effectively improved by appropriate application of nitrogen so that the photosynthetic capability and the dry matter accumulation of tobacco could be promoted, but over-application of nitrogen had a negative influence on photosynthetic efficiency. With increasing the rate of nitrogen, the activity of nitrate reductase increased and reached the maximum at 40 days after transplant for N1, N2, and 60 days after transplant for N3, showing that the nitrogen metabolism was postponed. The results also showed that the contents of total nitrogen and nicotine increased with increasing the rate of shading and nitrogen, but the content of carbohydrate reduced. Generally, the nitrogen metabolism was likely stronger than the carbon metabolism. Light, nitrogen and their interactions had a significant influence on flue-cured tobacco. The combination of 70% of natural light intensity and proper application of nitrogen (N2, 3.5 g per pot in this experiment) could improve the photosynthetic capability and the balance of carbon-nitrogen metabolism, and increase the amount of dry matter accumulation and the quality in flue-cured tobacco.

RESEARCH ACTIVITIES

Cloning and Comparative Analysis of PURPLE ACID PHOSPHATASE 17 Gene Families in Brassica oleracea and Brassica rapa

LU Kun,ZHANG Kai,CHAI You-Rong,LU Jun-Xing,TANG Zhang-Lin

Acta Agron Sin. 2010, 36(3):  517-525.  doi:10.3724/SP.J.1006.2010.00517

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Two PAP17 genes were cloned from parent species of Brassica napus, B. oleracea and B. rapa, respectively. According to sequence similarity, PAP17 genes could be divided into two types, type I (BoPAP17-1 and BrPAP17-1) and type II (BoPAP17-2 and BrPAP17-2). Southern hybridization resulted in two bands both in B. oleracea and B. rapa, this is accordance with former cloning results. Phytogenetic and molecular evolution analysis indicated that PAP17 genes in Brassica species underwent purifying selection, and their deduced proteins are typical low molecular weight PAP proteins. Expression patterns of BoPAP17 and BrPAP17 genes were assayed by fluorescent quantitative PCR. The results revealed that PAP17 genes expressed in all nine tested tissues and organs, with the extremely high expression in flower and bud, and certain expression in seeds at different stages, implying these PAP17 genes most likely mobilize phosphorus reserves in plants, particularly during flower and bud development stages. Under phosphate starvation conditions, expression of BrPAP17-2 in seedling leaf was restrained, while that of BrPAP17 and BoPAP17 in seedling root and leaf was induced, the expression levels declined in the first 24 hours, and then continuously increased with the maximal levels between four days and eight days after treatment. After four days of Pi-resupply, their expression declined below un-induced basal levels. In comparison with seedling leaf, it seems that BrPAP17 and BoPAP17 showed stronger phosphate starvation induced expression in seedling root. These results thus suggested that PAP17 genes in B. oleracea and B. rapa may be involved in external phosphorus assimilation and transferring inorganic phosphate from root to other tissues or organs.

Gene Cloning and Sequence Analysis of γ-Gliadin Genes from Wheat Cultivar Shann 253

WANG Ming-Xia,GAO Xiang,CHEN Qi-Jiao,DONG Jian,ZHAO Wan-Chun,LI Yan-Liang et al

Acta Agron Sin. 2010, 36(3):  526-532.  doi:10.3724/SP.J.1006.2010.00526

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Eight γ-gliadin genes (GenBank accession number GQ87170 to GQ87177) were cloned from wheat (Triticum aestivum L.) cultivar Shaan 253 using three sets of specific primers, which were designed according to the known γ-gliadin gene family. All of the eight genes possess the typical structure of γ-gliadin subunits and many sequences of insent/indel in the repeat region. The deduced amino acid sequences showed that GQ871771 is a pseudogene, and nine cysteine residues existed in GQ871770, GQ871772, GQ871776, and GQ87177. Promoter sequences analysis indicated that there were six SNPs in endosperm box of GQ871770, GQ871772, GQ87174, and GQ871776, and two of them appeared in the GCN4-like motif. A model for endosperm box of storage protein with better representation was constructed using the online tool WebLogo3. Phylogenetic analysis also confirmed that the cloned sequences belong to γ-gliadin gene family.

Cytoplasmic Alkalization Mediates Exogenous Nitric Oxide-Induced Stomatal Closure in Vicia faba

ZHAO Shi-Ling,SUN Li-Rong,ZHANG Huan,MA Li-Ya,LU Bao-Shi,HAO Fu-Shun*

Acta Agron Sin. 2010, 36(3):  533-538.  doi:10.3724/SP.J.1006.2010.00533

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The effects of exogenous NO on changes in cytosolic pH of guard cells from epidermal fragments and of the pH on NO-induced stomatal closure in Vicia faba were investigated by using a pH specific fluorescence probe SNARF-1-AM and a confocal laser scanning microscopy. The results showed that treatments with 100 μmol L–1 sodium nitroprusside (SNP), a widely used NO donor, led to a significant increase in cytosolic pH (from 6.91 to 7.19, i.e. 0.28 pH unit) and stomatal closure in comparison with the control. Furthermore, SNP-induced cytosolic alkalization and stomatal closure were significantly inhibited by NO scavenger c-PTIO or weak acid butyrate, and promoted by the weak base benzylamine. Whereas two structural analogs of SNP such as Fe(II)CN and Fe(III)CN, which can not release NO, failed to induce guard cell cytoplasmic alkalization and stomatal closure. These findings suggest that cytoplasmic alkalization of guard cells mediates NO-induced stomatal closure. In addition, pH in guard cell vacuoles and cell walls was measured during the cytosolic alkalization. However, no significant changes of pH in the two regions were found, implying that the cytoplasmic protons of guard cells may not go into vacuoles or cell walls during the NO-promoted alkalization in Vicia faba.