Table of Content

16 October 2016, Volume 49 Issue 20

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CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Phenotype Analysis and Gene Mapping of Narrow and Rolling Leaf Mutant nrl4 in Rice (Oryza sativa L.)

LIANG Rong, QIN Ran, ZENG Dong-dong, ZHENG Xi, JIN Xiao-li, SHI Chun-hai

Scientia Agricultura Sinica. 2016, 49(20):  3863-3873.  doi:10.3864/j.issn.0578-1752.2016.20.001

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【Objective】 Rice leaf mutant was used to study molecular mechanisms of leaf traits, and to identify the related novel rolling genes in rice.【Method】 The mutant with narrow and rolling leaves was derived from the indica cultivar Zhenong34 induced by ethyl methylsulphonate (EMS), named nrl4. At heading stage, nrl4 and WT Zhenong 34 were randomly selected 10 strains to measure the main agronomic traits and chlorophyll content of nrl4 and WT were tested at the same time. The bulliform cells were observed and counted as well as the number of large and small veins in transverse section of blade under the Zeiss microscope. The leaf phenotype of the F₁ plants and F₂ population which derived from the crossing of nrl4 with Zhenong 34 were investigated and the segregation ratio of normal and rolling leaves were analyzed by chi-square test in the F₂ population. The F₂ population from crossing of nrl4 with Zhenong 104 was used for genetic analysis and gene fine mapping. Five genes in the located region were analyzed by gene quantitative expression. 【Result】 Morphological analysis showed that all leaves of mutant nrl4 were narrow and rolling. In addition, compared with wild type Zhenong 34, plant height, seed setting rate in main panicle and pigment content of mutant nrl4 were increased, as well as grain length of nrl4, but the width of grain was decreased. Leaf angles of functional leaves were all decreased leading to more erecting plant type. Statistical analysis suggested that the rolling leaf phenotype might be caused by the change of number and size of bulliform cells which especially existed at the adaxial side of blade; moreover, in accordance with reduced leaf blade width, leaves of nrl4 contain a decreased number of large veins and small veins. There were 6.0 small veins between two large veins on one side of main vein averagely in mutant nrl4 leaf while there were 4.5 in wild type Zhenong34. Genetic analysis indicated that the mutant trait was controlled by a single recessive gene, the gene nrl4 was located in a confined region of 53 kb flanked by two InDel makers 3M11103 and 3M1115 on the long arm of chromosome 3, where five annotated genes were predicted. Based on the result of sequencing, there was no mutation occurred in the gene sequence and promoter sequence of these predicted genes, but strong changes in gene expression pattern of LOC_Os03g19770 according to the real time quantitative PCR. These results are very valuable for further study on this gene. 【Conclusion】 The narrow leaves are related to reduced number of vasculars, moreover the rolling blade of mutant nrl4 mightresulted fromthe decreased area and number of bulliform cells. The mutant nrl4 is controlled by a single recessive nuclear gene, which is located on chromosome 3, between 3M11103 and 3M1115 with a physical distance of 53 kb. No nucleotide sequence mutation was found to occur in the gene sequence or the 5′UTR of all annotated genes, but the expression of LOC_Os03g19770 is strongly promoted in mutant nrl4, which is 17.5 times of wild type and it may be the candidate gene.

Research on Methodology of Maize Germplasm Development with Source of Hybrids by Using Marker-Assisted Selection

CONG Chun-sheng, LI Yong-xiang, LI Chun-hui, SHI Yun-su, SONG Yan-chun, ZHANG Deng-feng, LI Yu, WANG Tian-yu

Scientia Agricultura Sinica. 2016, 49(20):  3874-3885.  doi:10.3864/j.issn.0578-1752.2016.20.002

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【Objective】 Research on marker-assisted methodology of developing germplasm suited as female and male parents was of practical importance in improving utilization efficiency of this source and promoting the development of maize breeding, since single-cross hybrids are over 95% in maize production. 【Method】 A popcorn hybrid, "JB1", was used as the initial entry in the present study to obtain polymorphic simple sequence repeat (SSR) markers and determine the parents’ genotype by performing polymerase chain reaction (PCR) amplification with genome-wide SSR markers and native polyacrylamide gel electrophoresis (PAGE). Subsequently, marker-assisted selection was performed in progenies of F₂ and two later subsequent generations using the polymorphic markers distributed evenly on the ten chromosomes of maize. As a result, three groups of progenies were obtained, i.e. male parent-like, female parent-like and middle type. To evaluate the heterosis and the breeding value of these selected progenies, hybrid combinations were made by using incomplete diallel cross in F₄ among the progenies from the male-like and female-like group which selected by marker-assisted selection. Testcrosses were also made between three testers and the materials selected by marker-assisted selection and breeding experience in early generation, respectively. Yield experiments and comprehensive evaluation of 2 780 hybrids were then conducted in Shunyi of Beijing and Yuxian of Hebei. 【Result】 Marker-assisted selection significantly improved the selection efficiency in the separate communities. In the F₄ generation, the similarity to the male parent and the female parent of selected materials could be as high as 79.5% and 73.7%, respectively. The similarity to the parents was much higher than that under the condition of random selection. Genetic distance between the materials from the male-like group and the female-like group was large and the maximum was 86%. Phenotypic analysis showed that the average plot yield of hybrid combinations between the male-like group and the female-like group was higher than that of hybrid combinations within each of the two groups, and there were no combinations whose yield exceeded the control (the original hybrid, JB1). Meanwhile, the testcrosses generated from the male-like materials displayed stronger heterosis and had higher plot yields than those generated from the female-like materials and the materials selected by breeding experience. There also existed some hybrid combinations whose yields exceeded the control. 【Conclusion】The study proposed a technical route of SSR marker-assisted germplasm development based on maize hybrids which could be useful in promoting use efficiency of maize hybrids.

TILLAGE & CULTIVATION·PHYSIOLOGY & ECOLOGY

Enzyme Activities and Soil Nutrient Status Associated with Different Aggregate Fractions of Paddy Soils Fertilized with Returning Straw for 24 Years

LI Wei-tao, LI Zhong-pei, LIU Ming, JIANG Chun-yu, WU Meng, CHEN Xiao-fen

Scientia Agricultura Sinica. 2016, 49(20):  3886-3895.  doi:10.3864/j.issn.0578-1752.2016.20.003

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【Objective】Paddy soils in subtropical China derived from Quaternary red clay are generally deficient in available nutrients, and fertilizers have long been identified as dominant contributors to increase of crop production. Therefore, a long-term experiment was set up in a paddy field that used to be a wasteland earlier to study the distribution of nutrients and enzyme activities in water stable aggregates under long term application of fertilizers, and the findings will provide a sound basis for rational application of fertilizers to ensure sustainable crop production. 【Method】Soil samples were collected from a 24-year long-term field experiment, which was established in 1990 in the Yingtan Red Soil Ecological Experiment Station. The experiment included four treatments: CK (without fertilization), SM (straw application plus manure), NSM (straw application plus manure and nitrogen fertilizer), NPKSM (straw application plus manure, nitrogen, phosphorus and potassium fertilizers). Undisturbed bulk soils were separated into five aggregate-size classes (＞2 mm, 1-2 mm, 0.25-1 mm, 0.053-0.25 mm and ＜0.053 mm) by wet sieving. Three soil enzymes, invertase, urease, and phosphatase, in water-stable aggregates (WSA) and total nitrogen (TN), available phosphorus (AP), and soil organic C (SOC) were determined. 【Result】Application of fertilizers to infertile paddy soil significantly increased the percentage of macro aggregates (larger than 0.25 mm), decreased the percentage of micro aggregates (smaller than 0.25 mm), and increased the mean weight diameter (MWD) of WSA, which could improve soil structure. The combined application of straw, manure and inorganic fertilizer significantly increased the soil enzyme activities in each size fraction. NSM treatment had the largest impact on invertase and urease activities. NPKSM treatment had the most significant impact on acid phosphatase activity. Compared with the control, NSM treatment increased invertase activity in five size fractions by 20.3%-396.2%; urease increased by 58.6%-372.1%. In NPKSM treatment, acid phosphatase activity in five size fractions increased by 48.9%-94.5%. Compared with the control, SOC in each size fraction of NSM treatment increased by 31.6%-65.1%. Total N increased by 19.8%-51.9%. In NPKSM treatment, available phosphorus content in each of the size fractions increased by 7.4-10 times. Aggregated boosted trees (ABT) analysis showed that the relative influence of SOC on invertase was the largest, accounting for 40.6% of the variation; the composition of soil particle had the largest relative influence on the activity of urease, accounting for 44.9% of the variation. Soil AP had the largest contribution of 41% to the variation in the activity of acid phosphatase. Nonmetric multidimensional scaling (NMDS) analysis indicated that soil aggregates within NPKSM treatment differed from those in CK, SM and NSM treatments. However, SM and NSM treatments showed similar effect on soil fertility. 【Conclusion】The combined application of straw, manure, and inorganic fertilizer significantly increased the mean weight diameter (MWD) of WSA, the contents of SOC, TN, AP, and soil enzyme activities, which improved the soil structure and biological functions.

Effects of Lime Application Rates on Soil Acidity and Barley Seeding Growth in Acidic Soils

HU Min, XIANG Yong-sheng, LU Jian-wei

Scientia Agricultura Sinica. 2016, 49(20):  3896-3903.  doi:10.3864/j.issn.0578-1752.2016.20.004

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【Objective】The aim of this study was to assess the effects of lime on soil acidity and barley seedling growth in an acidic soil (pH 3.9), and to determine the suitable dosage of lime and thus providing scientific proof for acidic soil improvement and reasonable application rate. 【Method】 The trial was conducted at the experimental base of Huazhong Agricultural University. Soil incubation and pot experiments were used. The Ca(OH)₂ titration method was employed to estimate the lime requirement, and six treatments were designed: no lime, 0.3 g·kg^-1, 0.9 g·kg^-1, 1.8 g·kg^-1, 2.4 g·kg^-1 and 4.8 g·kg^-1 lime application rates. Samples were collected on 10, 20, 30, 40, 50, 60, 70 and 90 days after incubation to monitor the dynamic changes of soil pH, soil exchangeable acidity, exchangeable H⁺ and exchangeable Al³⁺. Barley was sowed on the 90th day and harvested after 2 weeks to assess the effects of liming rates on barley biomass, morphological parameters, and root activity. 【Result】 Soil incubation results demonstrated that lime application significantly improved soil pH, however the content of exchangeable acidity and exchangeable Al³⁺ were decreased significantly during the first 30 days. The content of potential acid decreased gradually with the increasing dosage of lime, therefore the content of soil exchangeable acid, exchangeable H⁺ and exchangeable Al³⁺ were zero in the treatment of 4.8 g·kg^-1. From then on, due to the influences of soil buffer, low lime dosages (< 1.8 g·kg^-1) showed no obvious effects on improving soil pH but significantly reduced soil potential acidity. The results of pot experiment showed that application of lime significantly enhanced barley growth through improving plant height, biomass and root development. Barely seedlings plant height, dry mass, total root length and surface area, and root activity improved with the lime application rate from 0 to 1.8 g·kg^-1. Whereas, lime input decreased root average diameter. Barley growth was inhibited significantly when lime application rate was above 1.8 g·kg^-1. The root activity in the treatment of 4.8 g·kg^-1 was lower than that of 0.9 g·kg^-1, the excess of lime application retarded the root growth. Thus, the optimum liming rate was 1.8 g·kg^-1, which agreed with the lime requirement calculated using the Ca(OH)₂ titration method. 【Conclusion】 Lime application is capable of neutralizing soil acidity and promoting barley seedling growth. Under the conditions of this study (soil pH 3.9), the appropriate liming rate was 1.8 g·kg^-1 (4 t·hm^-2). This rate was consistent with that from the Ca (OH)₂ titration method, confirming that the approach adopted in this study is appropriate for determining the lime application rate.

Effect of Technique of Ridge Film Mulching and Furrow Seeding on Soil Erosion and Crop Yield on Sloping Farmland in Western Liaoning

XIAO Ji-bing, SUN Zhan-xiang, JIANG Chun-guang, ZHENG Jia-ming, LIU Yang, YANG Ning, FENG Liang-shan, BAI Wei

Scientia Agricultura Sinica. 2016, 49(20):  3904-3917.  doi:10.3864/j.issn.0578-1752.2016.20.005

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【Objective】Western Liaoning is not only short and great variation in precipitation resource, but soil erosion in hilly area is mostly serious in Liaoning province. The sloping farmland is the main source of water and soil loss. Ridge film mulching and furrow seeding could collect natural precipitation and reduce soil erosion effectively, improve water and soil resources utilization availability. The purpose of this study is to make full use of the limited natural rainfall, protect water and soil resources of dry farming area and promote high and stable yield of crops.【Method】Two factors split plot design was used to examine the effects of different farming types on soil erosion, soil water and yield in sloping farmland and the study was carried out by runoff plots under natural rainfall from 2012 to 2015 in dry farming experimental area in Liaoning Academy of Agricultural Sciences at Fuxin. The runoff plot was 15 m and 6 m in length and width and the area was 60 m². The main plot was a sloping land with slopes of 5°and 10°, and split plot was crop planting patterns, the designed patterns were traditional cultivation(CK, contour ridge and furrow planting, ridge and furrow were uncovered), ridge film mulching and furrow straw mulching(T1), ridge film mulching and furrow seeding(furrow was uncovered, T2). The study was repeated three times. Ridges and furrows of ridge film mulching and furrow seeding were in opposite arrangement, furrow width was 60 cm and ridge width was 40 cm, ridge height was 10-12 cm. The ridge was a rain collecting area. The furrow, which was planted with two lines of crop, was a planting area. The experimental crops were millet and maize and rotated.【Result】The ridge film mulching and furrow straw mulching and ridge film mulching and furrow seeding could effectively reduce and prevent water and soil erosion. In four years, the total runoff and total erosion amount of the CK were 512.7 m³·hm^-2, 4 561.3 kg·hm^-2and T1, T2 did not occur soil erosion under five-degree slope. The total runoff and total erosion amount of T1 were reduced by 81.71% and 96.17% and that of T2 were reduced by 56.92% and 95.15% compared with the CK, and that of T1 were reduced by 57.54% and 21.05% compared with T2, T1 and T2 would reduce the amount of lost nutrients in sediment compared with the CK under ten-degree slope. The results showed that runoff and erosion amount increased in accordance with increasing of gradient and the effect of anti-erosion of T1, T2 would weaken along with slope increasing. There was no significant difference between the five-degree slope and ten-degree on four years' average soil water, which had a reducing trend with slope increasing. The effect of T1, T2 on mean soil water reached very significant level as a whole compared with CK and mean soil water of T1, T2 was 1.68 and 1.45 percentage points higher than that of CK. The effect of T1, T2 on increasing moisture reached very significant level compared with CK to millet regardless of high flow year or dry year and that of T1, T2 on increasing moisture to maize was not significant in normal year and very significant in special dry year. There was no significant difference between five-degree slope and ten-degree slope in four years' average yield, which had a reducing trend with slope increasing. The effect of T1 on the average production reached very significant level increasing by 25.59% and 10.68% compared with CK and T2, and there was a significant difference between T2 and CK and the average production of T2 increased by 13.47% compared with CK. The effect of yield increasing of T2 was insignificant compared with CK and that of T1, which increased by 24.75% and 74.58%, was significant compared with CK for millet in 2012, 2014. The effect of T1, which the yield of maize increased by 11.29% and 54.39%, and T2, which increased by 5.05% and 51.81%, on the production reached very significant level compared with CK in 2013 and 2015. The effects of yield increasing of ridge film mulching and furrow straw mulching (millet, maize) and ridge film mulching and furrow seeding (maize) were particularly significant in dry year. 【Conclusion】 The effect of micro-rainwater-collecting planting mode by ridge film mulching and furrow seeding in dry sloping farmland on rainwater harvesting, water storage, soil moisture conservation and anti-erosion was definite after 4 years of study in semi-arid region in Liaoxi area. The technique could mitigate the negative impact of drought and soil erosion on crop growth and enriched the theoretical basis of the rain-harvesting agriculture of dry farming in the area. By application and dissemination of the technique, it could improve soil and water resources utilization, protect quality of cultivated land and promote overall grain production capability on sloping farmland and facilitate stable and high yield of crop and healthy and sustainable development of dry farming in the area. It could be seen that the ridge film mulching and furrow straw mulching would be suitable for dry sloping land in Western Liaoning from the point of anti-erosion, catchment, increasing soil moisture, increasing production. The study has filled the gaps in the study of this subject in western Liaoning and provided important references for development of the rain-harvesting agriculture of dry farming in the north of China.

PLANT PROTECTION

Strain Composition of Potato virus Y in Fujian Province Detected with the Concatenated Sequence Approach

SHEN Lin-lin, ZOU Wen-chao, GAO Fang-luan, ZHAN Jia-sui

Scientia Agricultura Sinica. 2016, 49(20):  3918-3926.  doi:10.3864/j.issn.0578-1752.2016.20.006

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【Objective】Potato virus Y (PVY) is one of the most destructive pathogens constraining sustainable development of potato industry. The objective of this study is to develop a fast, easy-to use and accurate approach to timely detect PVY strains and apply the approach to investigate the occurrence, distribution and composition of PVY strains in Fujian province. 【Method】ELISA method was used to confirm the presence of virus in the PVY-alike leaf samples randomly collected from Changle and Fuqing cities in Fujian province. Three pairs of degenerate primers designed from the conserved regions of P1, VPg and CP genes in the reference PVY sequences downloaded from GenBank were used to amplify the positive samples by ELISA test. Nucleotide identity and recombination events between isolates from Fujian and strains downloaded from GenBank were evaluated and phylogenetic tree was reconstructed using concatenated sequences of the three genes. 【Result】ELISA confirmed that 13 out of the 17 samples collected from the two potato producing areas were infected with PVY. RT-PCR amplifications of all 13 ELISA-positive samples generated the expected sizes of fragments corresponding to the three genes. Individual gene analyses showed that P1, VPg and CP sequences in Fujian isolates shared 72%-99%, 85%-99% and 88%-99% nucleotide identity with the reference strains, respectively. Concatenated sequence analysis showed that FQ01 and FQ08 isolates shared the highest sequence identity with PVY^N-Wi and PVY^E, respectively; CL01, CL02, CL05 and CL13 isolates shared the highest sequence identity with PVYN^TN-NW (SYR-I), whereas CL03, CL04, FQ02, FQ06, FQ09, FQ11 and CL12 shared the highest sequence identity with PVY^NTN-NW (SYR-II). Recombination signals were identified in P1 and VPg genes but not in CP of PVY, which isolated from Changle and Fuqing cities in Fujian province. Further analyses showed that in Changle City, P1 gene was composed of two lineages, with 60% being N lineage and 40% being N×O lineage. All VPg genes were N×O lineage and all CP genes were classified as O lineage. Similarly in Fuqing city, P1 gene was consisted of two lineages, with 25% being N lineage and 75% being N×O lineage; 87.5% VPg genes were N×O lineage and 12.5% was O lineage; and all except one (which was classified as N lineage) CP genes were O lineage. Phylogenetic tree revealed that isolates CL01, CL02, CL05 and CL13 were grouped with PVY^NTN-NW (SYR-I) and isolates CL03, CL04, FQ02, FQ06, FQ09, FQ11 and CL12 were clustered with the PVY^NTN-NW (SYR-II), whereas isolate FQ01 was grouped with PV^YN-Wi and isolate FQ08 was clustered with the PVY^E, respectively. These results indicated that evolutionarily, CL01, CL02, CL05 and CL13 are closer to PVY^NTN-NW (SYR-I); CL03, CL04, FQ02, FQ06, FQ09, FQ11 and CL12 are closer to PVY^NTN-NW (SYR-II), FQ01 is closer to PVY^N-Wi and FQ08 is closer to PVY^E, respectively. 【Conclusion】PVY occurs frequently in Changle and Fuqing cities in Fujian province and recombinant strains, particularly PVY^NTN-NW are dominant in the PVY isolates from the province.

Construction and Transformation of RNAi Vector for Citrus tristeza virus Gene p23

LI Fang, DENG Zi-niu, ZHAO Ya, LI Da-zhi, DAI Su-ming

Scientia Agricultura Sinica. 2016, 49(20):  3927-3933.  doi:10.3864/j.issn.0578-1752.2016.20.007

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【Objective】 The objective of this study is to construct RNAi vector containing p23 gene of Citrus tristeza virus (CTV), and obtain transgenic orange plants with virus resistance. 【Method】 Based on the pathogen-derived resistance and the CTV genome sequences published by NCBI, two specific conserved fragments of p23 with different lengths were cloned. Two segments and vector pBI 121 were double-digested and connected for RNAi construction. Subsequently, to initially estimate the antiviral feasibility, the Mexican lime (CTV indicator plant) leaves were injected with Agrobacterium contained the RNAi vector for transient expression, and observed using GUS histochemical staining method. The leaves were inoculated with CTV T36 isolate, and detected by enzyme-linked immunosorbent assay (ELISA). The RNA of leaves was also extracted and reverse transcript to cDNA. Quantitative real-time PCR (q-PCR) was performed to observe the CTV p20 gene expression which could reflect the virus in hosts. The RNAi vector was also transferred into the epicotyl stem of ‘DA HONG’ sweet orange seedlings via Agrobacterium-mediated transformation. Resistant buds were engrafted onto Carrizo Citrange in vitro seedlings. DNA extracted from the ‘DA HONG’ sweet orange leaves was used as the PCR template to identify the transgenic plants. Plants containing the target gene were re-engrafted onto sour orange seedlings and stored in the greenhouse. The expression of the p23 within the transgenic plants was evaluated by q-PCR. The skin buds of CTV T36 isolate hosts were collected and inoculated onto the transgenic sweet orange. The leaves from the sprouted branch tips were collected and analyzed the pathogen resistance capability with the same method that the transient expression leaves detected. Plants without detectable virus infection after the first inoculation were also inspected and analyzed by the same manner in the second round. 【Result】 Long (513 bp) and short (291 bp) fragments of the p23 were cloned. These p23 fragments are then cloned into the pBI121 vector, named p23-RNAi. This p23-RNAi vector was then delivered in the Mexican lime leaves using the Agrobacterium-mediated transient expression assay. The leaves were identified based on the presence of blue stains after GUS staining, indicating that the Agrobacterium contained vector p23-RNAi may induce transient expression in Mexican lime leaves. On the 15th and 30th day after the CTV inoculation, the ELISA detection results for the transgenic Mexican lime leaves in p23-RNAi plants were all negative, whereas the q-PCR detection results showed that the accumulation level of p20 expression was significantly lower than that of the control plants. It indicated that the transient expression of p23-RNAi may, in a defined period, inhibit the CTV infection. Introduction of p23-RNAi via Agrobacterium-mediated genetic transformation led to the production of resistant buds for the ‘DA HONG’ sweet orange, and PCR amplification confirmed a total of seven transgenic plants. The expression of the p23 in all seven transgenic plants was further confirmed by q-PCR amplification, and the gene expression level exhibited a certain degree of difference, with expression level in plant E being the highest, followed by C, F, H, A, B, and G. After inoculation with CTV, the level of expression of p20 varied among these seven transgenic plants, with plant A having the highest level of p20 expression, followed by G, F, E, B, H and C. The transgenic plants showed higher pathogen resistance, albeit to different degrees, when compared to the control plants. However, the virus resistance degrees in the transgenic plants were not closely related to the expression levels of the exogenous gene. For example, plant E, which had the highest expression level of the exogenous gene, did not exhibit powerful CTV resistance. By contrast, the transgenic plant C displayed complete resistance after the two rounds of virus inoculation. 【Conclusion】The p23-RNAi construct that generated confers plant disease resistance to CTV. Transient expression assay can be applied for the high efficiency identification of resistance and screening for high efficiency RNAi vectors.

SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT

Nitrogen of Organic Manure Replacing Chemical Nitrogenous Fertilizer Improve Maize Yield and Nitrogen Uptake and Utilization Efficiency

XIE Jun, ZHAO Ya-nan, CHEN Xuan-jing, LI Dan-ping, XU Chun-li, WANG Ke, ZHANG Yue-qiang, SHI Xiao-jun

Scientia Agricultura Sinica. 2016, 49(20):  3934-3943.  doi:10.3864/j.issn.0578-1752.2016.20.008

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【Objective】 Replacing chemical fertilizer with organic manure is an important way to achieve fertilizer zero increase in China. Eight years location experiments of replacing chemical fertilizer with organic manure were carried out to study maize productivity and nitrogen uptake and nitrogen utilization efficiency in purple soil, so as to provide scientific evidence about rational utilization organic resources and adjust maize fertilization pattern.【Method】In 8 years location experiments, five different fertilization patterns were designed, including CK (non-N fertilization), FP (farmers’ practice), OP (optimal chemical fertilization), MF (organic manure replacing 50% chemical N) and OM (organic manure replacing 100% chemical N), on the variation of maize yield and biomass and nitrogen uptake and nitrogen utilization efficiency.【Result】Replacing partial chemical with organic manure increased maize yield and biomass significantly. Compared with FP and OM and OP treatments, replacing 50% chemical N with organic manure increased maize yield by 13.7% and 13.5% and 12.5%, respectively; it also increased maize biomass by 11.3% and 7.0% and 8.6%, respectively. Compared to CK, the other four treatments decreased the coefficient of variation (CV) and increased the SYI and HI with the greatest change in MF. Organic manure replacing partial chemical N promoted N uptake in maize, and transferred more N to grain. Compared to OP and OM treatments, replacing 50% chemical N with organic manure increased grain N uptake by 7.0% and 29.6%. RE_N increase 2.5% and 26.5%, respectively. Compare to OP treatment, PFP_N and NHI and FCR_N in MF increased by 6.2 kg·kg^-1 and 3.5% and 6.3%, respectively; Compared to OM treatment, increased by 6.6 kg·kg^-1 and 0.8% and 5.8%, respectively. There was a significant difference in N demand for each ton of grain yield among treatments, with 9.4 and 10.8 kg in OP and MF treatments, respectively, which was less than that in FP and OM treatments (14.5 and 12.9 kg). Therefore, OP and MF could improve the N efficiency for grain production. 【Conclusion】 Replacing 50% chemical N with organic manure significantly increased maize economic yield and biomass, also increased yield stability and sustainability; It promoted N uptake in maize and transferred more N to grain, thus increasing N efficiency. So partial chemical N is replaced by organic manure is a rational fertilization pattern to increase maize yield and improve maize yield stability and improve N efficiency in Southwest China.

Effect of Mulching and Nitrogen Fertilizer on Maize Yield，Distribution and Fate of Nitrogen in Root Layer

WANG Xiu-kang, XING Ying-ying, LI Zhan-bin

Scientia Agricultura Sinica. 2016, 49(20):  3944-3957.  doi:10.3864/j.issn.0578-1752.2016.20.009

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【Objective】 The objective of this study is to investigate the effect of mulching and nitrogen fertilizer application rate on root zoon nitrogen distribution, fate and maize yield for rational application of fertilizer and agricultural sustainable development in the Loess Plateau of Northwest China. 【Method】 Six treatments were designed and applied: (1) A flat plot with no basal fertilizer, no top dressing and no mulching (CK); (2) Plastic film mulching with no basal fertilizer and no top dressing (MN0); (3) Basal N (80 kg·hm^-2) and P (80 kg·hm^-2) with no top dressing and no mulching (BN1); (4) Plastic film mulching and basal N (80 kg·hm^-2) and P (80 kg·hm^-2) with no top dressing (MN1); (5) Basal N (80 kg·hm^-2) and P (80 kg·hm^-2) and top dressing N (80 kg·hm^-2) with no mulching (BN2); and (6) Plastic film mulching with basal N (80 kg·hm^-2) and P (80 kg·hm^-2) fertilizer and top dressing N (80 kg·hm^-2) (MN2). The effect of mulching and nitrogen fertilizer on maize yield, soil water content, soil nitrate-N distribution and the uptake of nitrogen of above-ground part of maize were analyzed.【Result】 The shoot dry matter accumulation of maize was increased with the increase of growth period, and the rate of dry matter accumulation was also increased. The shoot dry matter accumulation showed mainly the MN2＞BN2＞MN1＞BN1＞CK＞MN0. The more the shoot dry matter accumulated, the higher the grain yield of maize. Mulching and nitrogen fertilizer application had a significant effect on grain yield of maize. The yields of BN1 and MN1 treatments were 31.41% and 38.33% higher than the CK treatment, and the yields of BN2 and MN2 treatments were 49.89% and 79.06% higher than the CK treatment in 2012. Mulching was significantly increased the soil water content, the soil water content was increased first, and then reduced. The soil nitrate-N content decreased with the advance of growing period in no nitrogen fertilizer treatment. The upper soil nitrate-N content was slightly larger than subsoil layer, and the difference in quantity was decreased with the advance of growing period. Mulching had the function of slow the migration of soil nitrate-N move to sub soil layer in basal and top-dressing nitrogen fertilizer treatment. The nitrogen uptake rate of above ground dry matter accumulation was positively correlated with the level of nitrogen fertilizer. Mulching and nitrogen fertilizer application had a significant effect on the nitrogen uptake rate. There was no significant difference in nitrogen uptake rate in no-fertilizer treatment. The fate of nitrogen in mulching treatment showed mainly the nitrogen uptake rate＞nitrogen residual＞nitrogen loss rate. Nitrogen application significantly improved the nitrogen recovery rate, which was showed mainly that the MN2＞BN2＞MN1＞BN1 in both years.【Conclusion】The MN2 treatment significantly increased the maize yield and soil water content in top soil layers, and reduced the speed of nitrate-N from top soil layers to subsoil layers. Meanwhile, the MN2 treatment also reduced the nitrogen loss rate and improved the nitrogen use efficiency and nitrogen uptake rate.

TILLAGE & CULTIVATION·PHYSIOLOGY & ECOLOGY

Synergistic Effects of Water-Saving Irrigation, Polymer-Coated Nitrogen Fertilizer and Urease/Nitrification Inhibitor on Mitigation of Greenhouse Gas Emissions from the Double Rice Cropping System

LI Jian-ling, LI Yu-e, ZHOU Shou-hua, SU Rong-rui, WAN Yun-fan, WANG Bin, CAI Wei-wei, GUO Chen, QIN Xiao-bo, GAO Qing-zhu, LIU Shuo

Scientia Agricultura Sinica. 2016, 49(20):  3958-3967.  doi:10.3864/j.issn.0578-1752.2016.20.010

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【Objective】Optimization of water and nitrogen management measures has a great significance for rice yield improvement and greenhouse gas emission reduction in paddy fields. To establish water and fertilization management regimes with effects on yield promotion and greenhouse gas mitigation, a new water-saving irrigation technique, “thin and shallow alternate wetting drying”, was investigated in the double rice cropping system. Synergistic effects of water-saving irrigation and new types of nitrogen fertilizer on rice yield and greenhouse gas emissions were evaluated. 【Method】The study focused on double rice cropping system in the Jianghan Plain, Hubei province, Central China. Greenhouse gas emissions were observed from four different treatments: U＋CI: urea with conventional traditional irrigation, as the control (CK); U＋SI: urea with “thin and shallow alternate wetting drying” water-saving irrigation; CRU＋SI: polymer-coated urea with “thin and shallow alternate wetting drying” water-saving irrigation; NU＋HQ＋SI: nitrapyrin crystal urea with hydroquinone and “thin and shallow alternate wetting drying” water-saving irrigation. Measurements were taken using the automatic static chamber-GC (gas chromatography) method. CH₄ and N₂O emissions, and total CO₂-eq (CH₄＋N₂O, on a 100a horizon) of each treatment were analyzed. Rice yield per plot and greenhouse gas intensity (GHGI) were calculated after harvesting. 【Result】The “thin and shallow alternate wetting drying” water-saving irrigation technique diminished CH₄ emission fluxes during the early and late rice seasons, especially at the reproductive stage, resulted in lower CH₄ emissions for U＋SI compared to U＋CI (P<0.01). The reduction in CH₄ emissions in the late rice season was greater than in the early rice season. By using water-saving irrigation techniques, pronounced differences in CH₄ emissions were identified among polymer-coated urea, nitrapyrin crystal urea with hydroquinone and urea treatments. Total CH₄ emissions during two rice seasons from CRU＋SI and NU＋HQ＋SI were 60% and 73% of emissions from the U＋SI treatment, respectively. “Thin and shallow alternate wetting drying” water-saving irrigation increased N₂O emissions in paddy fields. Compared to U＋CI, the U＋SI treatment significantly increased N₂O emissions in the early and late rice seasons by 34% and 39%, respectively (P<0.05). Compared to urea, N₂O emissions from nitrapyrin crystal urea with hydroquinone and polymer-coated urea treatments were decreased. The effect of nitrapyrin crystal urea with hydroquinone on the control of N₂O emissions was superior to that of the other two nitrogen fertilizers. A trade-off was identified between CH₄ and N₂O emissions under “thin and shallow alternate wetting drying” irrigation, but the effect on the reduction of CH₄ emissions was greater than the increase in N₂O emissions. Overall, “thin and shallow alternate wetting drying” irrigation can reduce total greenhouse gas emissions, with the strength of the mitigation effect depending on the type of nitrogen fertilizer used. Polymer-coated urea had the greatest mitigation effect, reducing total CO_2-eq emissions by 49%, followed by nitrapyrin crystal urea with hydroquinone (46%) and urea (28%). Furthermore, polymer-coated urea and nitrapyrin crystal urea with hydroquinone were more beneficial for rice yield promotion and decreased greenhouse gas intensity of rice production. 【Conclusion】The results suggest that “thin and shallow alternate wetting drying” water-saving irrigation has good effects on yield maintenance and GHG abatement. The combined application of water-saving irrigation and polymer-coated urea or nitrapyrin crystal urea with hydroquinone could further increase rice yield and reduce greenhouse gas emissions. Thus, these water and nitrogen management measures deserved wider extension in order to simultaneously increase grain yield and decrease global warming effects.

SOIL & FERTILIZER·WATER-SAVING IRRIGATION·AGROECOLOGY & ENVIRONMENT

Dissolved Methane Concentration and Diffusion Flux in Agricultural Watershed of Subtropics

ZHANG Yu, LI Yue, QIN Xiao-bo, KONG Fan-long, XI Min, LI Yu’e

Scientia Agricultura Sinica. 2016, 49(20):  3968-3980.  doi:10.3864/j.issn.0578-1752.2016.20.011

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【Objective】The objective of this study is to investigate the regulation of dissolved CH₄ and flux diffused from river water and its influencing factors. 【Method】A one year period of (from April 2014 to April 2015) monitoring was conducted in Tuojia watershed of Xiangjiang river, which is located in the red soil hilly area of subtropical China. The double-layer-diffusion model was used to measure the diffusion of CH₄ flux from river water and the influencing factors of water were monitored by a portable multi-parameter meter. Four reaches river of Tuojia watershed with 3 streams (up stream, middle stream and down stream) each were employed in this study. 【Result】The results indicated that the annual dissolved CH₄ concentration and diffusion flux of CH₄ from Tuojia river varied widely from 0.03 to 2.23(0.61±0.43) μmol·L^-1 and from 1.71 to 290.08(63.36±50.76) μgC·m^-2·h^-1, respectively. Tuojia river expressed as the net source of atmospheric CH₄. There was a significant spatial and temporal difference both in the CH₄ concentration and flux between the 4 reaches. Generally speaking, the CH₄ flux from down stream were greater than the up stream, and the same as the CH₄ concentration (S₄＞S₃＞S₂＞S₁). And the temporal variation of CH₄ concentration and flux between the 4 reaches was also significant. Spring period (from April 2014 to Jun 2014) showed the highest CH₄ concentration (0.74±0.41μmol·L^-1) and the highest flux (93.58±65.24μgC·m^-2·h^-1), and winter period (from Dec 2014 to April 2015) showed the smallest CH₄ concentration(0.53±0.38 μmol·L^-1) and the smallest flux (50.79±33.03 μgC·m^-2·h^-1). By correlation analysis, it was found that, on the one hand, the water dissolved oxygen(DO: 3.49-12.79 (7.90±1.78) mg·L^-1) appeared a significant negative correlation with CH₄ concentration(r=-0.39, P＜0.001) and CH₄ flux(r=-0.36, P＜0.001), while the dissolved organic carbon (DOC: 0.92-7.38 (2.99±1.25) mg·L^-1) showed a positive correlation with CH₄ concentration(r=0.50, P＜0.001) and CH₄ flux(r=0.44, P＜0.001), all of them were the dominant factors that influence the CH₄ concentration and CH₄ flux of Tuojia River, on the other hand, dissolved inorganic N (NH₄⁺-N: 0.02-4.37 (1.26±1.03) mg·L^-1, NO₃^--N: 0.24-2.66 (1.43±0.55) mg·L^-1) concentration, salinity(represented by electrical conductivity EC: 50.36-248.43 (138.37±47.54) μS·cm^-1) appeared a positive correlation with CH₄ concentration and CH₄ flux. Additionally, water pH value (5.89-8.54(6.82±0.31)) showed a positive relationship with stream CH₄ concentration (r=0.20, P＜0.05), but not a significant correlation with CH₄ flux. The dissolved CH₄ in river water was produced from river sediment by methanogenesis, and then diffused from water to atmosphere.【Conclusion】The results of this study indicate that waste and sewage produced by agricultural non-point source pollution, livestock breeding and human activities are the main reasons that leading to the increase of river pollution loading and the decrease of water DO, which give raises to more stream CH₄ transportation, made river as an important potential CH₄ source.

HORTICULTURE

Changes in Soluble Sugars and Organic Acids of Xinjiang Apricot During Fruit Development and Ripening

ZHENG Hui-wen, ZHANG Qiu-yun, LI Wen-hui, ZHANG Shi-kui, XI Wan-peng

Scientia Agricultura Sinica. 2016, 49(20):  3981-3992.  doi:10.3864/j.issn.0578-1752.2016.20.012

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【Objective】The aim of this study was to characterize the content and composition of soluble sugars and organic acids in Xinjiang apricot fruit and to reveal the change rule.【Method】In this study, the soluble sugars and organic acids in peels and pulps of five Xinjiang apricot cultivars were determined by high performance liquid chromatography (HPLC) during fruit development and ripening. 【Results】 Three sugars were identified from Xinjiang apricot fruit, including sucrose, glucose and fructose. Sucrose and glucose are the predominant soluble sugars in Xinjiang apricot fruit. At the full-ripe stage, the ratio of sucrose to total sugar was  60.7%-79.1% in peels and 65.5%-82.4% in pulps, whereas glucose occupied 13.5%-34.7% of total sugar in peels and 8.2%-25.9% in pulps, respectively. Fructose only made up 4.6%-10.6% and 6.5%-10.7% of total sugar in peels and pulps respectively. Three individual sugars identified and total sugar increased significantly throughout the whole development period. The ratio of glucose to total sugar decreased from 79.4% to 13.5% in peels and 74.1% to 8.2% in pulps, respectively. Conversely, the ratio of sucrose to total sugar increased from 11.0% to 79.1% in peels and 11.0% to 82.4% in pulps. A total of six organic acids, including malic acid, quinic acid, citric acid, tartaric acid, oxalic acid and fumaric acid, were identified from Xinjiang apricot. Malic acid, quinic acid and citric acid were the dominant organic acids in ripe fruit, occupying 94.6%-98.3% of total organic acid. During fruit development and ripening, the contents of malic acid and oxalic acid decreased dramatically, while the contents of citric acid, quinic acid and fumaric acid increased significantly, no clearly consistent trend was observed for tartaric acid. The content of total acid in fruit increased during early development (from fruitlet to enlargement stage), but decreased rapidly during fruit ripening (from turning to full-ripe stage). Though malic acid predominated in fruit throughout the whole development period, different accumulation patterns for organic acids were observed in all cultivars studied. The patterns were divided into two types: malic acid and quinic acid dominated, malic acid and citric acid dominated, or only malic acid dominated during early development period, yet all types changed to malic acid, quinic acid and citric acid dominated during ripening. However, the ratio of malic acid, quinic acid or citric acid to total organic acid differed significantly in cultivars tested during fruit ripening. The ratio order in ‘Kuerletuoyong’ (KE), ‘Akeyaleke’ (AK) and ‘Kezijianali’ (KZ) was citric acid > malic acid > quinic acid, the ratio in ‘Suogejianali’ (SG) was quinic acid > malic acid > citric acid, the ratio in ‘Sulian No. 2’ (SL) was that all three organic acids appeared to be largely equal. No significant differences were found in soluble sugars and organic acid content, composition or accumulation pattern between peels and pulps. 【Conclusion】Xinjiang apricot presented an obvious chemical changing trend of soluble sugars and organic acids accumulation during fruit development and ripening. The sugar accumulation pattern changed from glucose dominated to sucrose dominated, while the organic acids accumulation pattern changed from two or one of malic acid, quinic acid and citric acid to the accumulation of the three organic acids all together. Thus, this pattern may play an important role in sweetness, acidity and flavor determination for Xinjiang apricot fruit.

Mineral Element Contents of Four Main Fruits from Liaoning Province and Their Dietary Exposure Assessment

KUANG Li-xue, NIE Ji-yun, LI Zhi-xia, WU Yong-long, YAN Zhen, CHENG Yang, GUAN Di-kai

Scientia Agricultura Sinica. 2016, 49(20):  3993-4003.  doi:10.3864/j.issn.0578-1752.2016.20.013

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【Objective】A study of detection and dietary exposure assessment of main mineral element contents of 4 dominant fruits from Liaoning province (including apple, pear, grape and peach) was carried out for the purpose of providing references to fruit production and consumptions. 【Method】 A total of 140 samples of the 4 fruits were collected from the main producing areas，including Dalian, Anshan, Yingkou, Jinzhou, Chaoyang, Huludao and Suizhong. The contents of 10 mineral elements were detected by ICP-AES and the correlation was analyzed by SPSS17.0 statistic software. Daily intake assessments of mineral elements from the 4 fruits among different populations were conducted and analyzed using recommended nutrient intake (RNI) or adequate intake (AI) and tolerable upper intake level (UL), respectively. Total diet risk assessment for adult was analyzed with residents balanced diet pagoda and food composition table. 【Result】 The average contents of 10 mineral elements were followed by the order of K (367.68 mg·kg^-1)＞P (122.65 mg·kg^-1)＞Mg (78.93 mg·kg^-1)＞Ca（76.36 mg·kg^-1）＞Na（12.37 mg·kg^-1）＞B（3.03 mg·kg^-1）＞Fe（2.86 mg·kg^-1）＞Cu（1.17 mg·kg^-1）＞Mn（1.02 mg·kg^-1）＞Zn（0.82 mg·kg^-1）and the range of coefficient of variation was from 27.80%（Mg）to 82.05%（Cu）. There was a significant correlation between K and Mg. Along with the growth of the age, the ratios of K to its AI was declined while other elements were showed down-up-down. The maximal ratio of Fe was 6.54% found in 4-7 ages and other elements were in 2-4 ages. In which, the percentages of RNI of Cu, Mg, P, K, Zn, Ca and Na were 83.30%, 11.01%, 8.57%, 8.43%, 4.25%, 2.46% and 0.39%, respectively. Results of dietary risk assessment showed that the average and the highest risk index of Cu in 2-4 ages were 12.50% and 37.02%, respectively. The risk indexes of other mineral elements were all far below 100% with the range of 0.33%（Zn）-13.40%（Mn）. Risk indexes of total diet for adults were followed by Mn（53.82%）＞Fe（49.93%）＞P（46.27%）＞Ca（35.23%）＞Zn（33.50%）＞Cu（28.61%）calculated by UL, which were all below 100%. 【Conclusion】 Contents of K, P, Mg and Ca were always higher than other trace elements in fruits. Synergistic effect plays a key role among different elements. Intake of 10 mineral elements from the above 4 fruits are safe to human health. The risk index of Cu is higher than other elements and should be paid a special attention. The daily dietary intakes of 10 mineral elements from total diet will not pose a health risk to adults.

STORAGE·FRESH-KEEPING·PROCESSING

Separation and Antioxidant Activity of Different Phenolic Compound Fractions from Litchi Pulp

DONG Li-hong, ZHANG Rui-fen, XIAO Juan, DENG Yuan-yuan, ZHANG Yan, LIU Lei, HUANG Fei, WEI Zhen-cheng, ZHANG Ming-wei

Scientia Agricultura Sinica. 2016, 49(20):  4004-4015.  doi:10.3864/j.issn.0578-1752.2016.20.014

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【Objective】The total phenolics, total flavoniods and tannin content and antioxidant activity of different phenolic compound fractions from litchi pulp were compared to clarify the effective phenolic compounds of antioxidant activity from litchi pulp. The results of research will provide a basis for revealing the main compounds from litchi pulp to human health.【Method】Litchi pulp polyphenol extracts were divided into four phenolic compound fractions (F1, F2, F3 and F4) by C18 silica gel column. Then the total phenolics, total flavonoids and tannin content and the compositions of free phenol of four compound fractions were determined. Also, the FRAP, DPPH, ORAC and CAA antioxidation indexes were adopted to evaluate the antioxidant capacities of four different compound fractions.【Result】The results showed that litchi pulp polyphenol extracts were well-divided into four phenolic compound fractions. Among them, the yield of F2 was as high as 36%, and others’ were 18.71%, 16.79% and 21.12%, respectively. Total phenolics, total flavonoids and tannin content of four phenolic compound fractions ranged from 218.86 to 499.78 mg GAE/g DW, 414.94 to 1 285.45 mg RE·g^-1 DW and 83.35 to 483.43 mg CE·g^-1 DW, respectively. Among the four compound fractions tested, F2 exhibited the highest total phenolics, total flavonoids and tannin content with their corresponding percentage contribution as high as 50.31%, 54.24% and 72.06% to litchi pulp polyphenol extracts, followed by F3, F4, and finally F1. Procyanidin B2 and epicatechin in F2, quercetin-3-O-rutinoside-7-O-α-L-rhamnoside in F3 and rutin in F4 were identified by HPLC. The results of antioxidation indexes showed that FRAP values of four compound fractions were F2＞F3＞F4＞F1. The IC₅₀ values of DPPH free radical scavenging were 27.00, 9.76, 19.41 and 16.25 μg·mL^-1, respectively, then F2 exhibited the strongest DPPH free radical scavenging ability with minimum IC₅₀ value among the four compound fractions, followed by F4, F3, and finally F1. Also, the ORAC values of them were F2＞F3＞F4＞F1 and the CAA values were F2＞F3＞F4, F1. Among them, F2 exhibited the highest ORAC and CAA values as 8.36 mmol TE·g^-1 DW and 190.71 μmol QE·g^-1 DW with the percentage contribution as high as 50.42% and 84.91% to total ORAC and CAA values of litchi pulp polyphenol extracts, respectively.【Conclusion】These results indicated that there were significant differences in total phenolics, total flavonoids and tannin content and antioxidant activity among the four phenolic compounds fractions from litchi pulp. The compositions of each phenolic compound fraction were also different. What’s more, F2 exhibited the highest yield, total phenolics, total flavonoids and tannin content with the best antioxidant capacities. Interestingly, the phenolics in F2 may be the most main active compounds of antioxidant activity from litchi pulp.

The Sensory Evaluation Methods of Production Process and Product of Lanzhou Hand-Extended Noodles

WEI Yi-min, XING Ya-nan, ZHANG Ying-quan, KONG Yan, LI Ming, ZHANG Bo, TANG Na

Scientia Agricultura Sinica. 2016, 49(20):  4016-4029.  doi:10.3864/j.issn.0578-1752.2016.20.015

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【Objective】 Experiment, verification and statistical analysis were combined in this study to establish a set of scientific and rational sensory evaluation attributes for Lanzhou hand-extended noodles (LZHEN), in order to provide methods for noodle quality evaluation, developing specific flour, and breeding special wheat varieties, also providing reference or case for the establishment of the traditional food sensory evaluation methods. 【Method】 Eight kinds of commercial wheat flours and two kinds of wheat flours with large differences in the quality premixed in a certain ratio were used as raw materials to make noodles by professional masters in laboratory. Using a preliminary method, masters evaluated the process procedure and panelists evaluated the cooked noodles. Then analyze, improve, and test the accuracy and precision of sensory evaluation methods on process procedure and the cooked noodles of LZHEN. 【Result】 The experiments of eight kinds of wheat flours processed by one master and six kinds of premixed wheat flours processed by three masters were made in triplicate. The results showed that the sensory evaluation methods established in this study had less significant difference in different evaluation times, fitting line, high accuracy, which means that the methods could be used to evaluate LZHEN. 【Conclusion】 By design, implementation and verification, the main quality attributes finalized for the process procedure and cooked noodles quality of LZHEN were as follow: 1) degree of mixing and hardness of dough, fracture and homogeneity of strips, slitting effort by hand and stickiness, dough color after resting and extending force were decided to assess the processing procedure quality attributes. 2) color, surface appearance, hardness, stickiness, elasticity, smoothness and flavor were decided to assess the cooked noodles quality attributes.

ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT

An Association Study of Positional and Functional Candidate Genes HMGA1, C6orf106 and ENSSSCG00000023160 with Leg Soundness in Pigs

ZHANG Xu-fei, HOU Li-juan, QIU Heng-qing, HUANG Lu-sheng, GUO Yuan-mei

Scientia Agricultura Sinica. 2016, 49(20):  4030-4039.  doi:10.3864/j.issn.0578-1752.2016.20.016

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【Objective】 The objective of this study is to develop a method to access the leg soundness through scoring the joint of limb bone and calculate the simple correlation coefficients among the scores for leg soundness in pigs. Furthermore, the association between three positional and functional candidate genes, namely HMGA1, C6orf106 and ENSSSCG0000023160, and leg soundness was also studied in F2, Laiwu, Erhualian, Sutai and DLY populations.【Method】The joint of five limb bones were scored according to the size and depth of rip on the joint surface and the worn-out degree of the joint. If the rip on the joint surface is very big and deep or the joint is seriously worn out, the joint is scored 1. On the other hand, if there is no rip on the joint surface and the joint doesn’t have any degree of worn-out, the joint is scored 5. Higher the joint score is, healthier the joint is and sounder the leg is. Based on the authors’ previous genome-wide association studies, three genes HMGA1, C6orf106 and ENSSSCG0000023160 were screened as positional and functional candidate genes to leg soundness in a 0.4 Mb region centered on the top SNP on SSC7. To search the polymorphic loci of the three genes in the F2 population, their DNA sequences were determined by a short-gun DNA sequence method. A total of 11 polymorphic loci were picked out according to their conservations among 6 species. The genotypes of 3 loci for HMGA1 and 3 loci for C6orf106 were determined using the Taqman method, and the genotypes of the other 5 loci for the third gene were inferred by genotype imputation just in the F2 population. At last, the GenABEL package of R was used to perform the association analysis between the loci with MAF＞0.05 and the traits.【Result】A total of 174 and 5 polymorphic loci were identified in C6orf106 and ENSSSCG00000023160 genes, respectively. Joint scores were positively correlated with each other and were negatively correlated with the length and weight of biceps brachii, but most of them had no correlation with toe, leg and gait scores. The male’s scapula joint score was significantly lower than the female’s, but arm shoulder joint score and focile hock joint score were significantly higher than the female’s corresponding joint score. In the F2 population, all of the three genes were associated with leg soundness, but ENSSSCG00000023160 was weaker than the other two genes, therefore it was excluded as a candidate gene to leg soundness and was not genotyped in the other 4 populations. In the Erhualian population, two loci g.2029C＞T and g.3155A＞G of HMGA1 were significantly associated with leg soundness, and the other SNP lacked the polymorphism. In the other 3 populations, all of the 3 SNPs of HMGA1 were deficiently polymorphic. Only the g.6953T＞C locus of C6orf106 had enough polymorphic in the Laiwu population, and it was associated with leg soundness. In the Sutai and DLY populations, only two loci g.2054T＞C and g.6953T＞C of C6orf106 were polymorphic, but none was associated with leg soundness.【Conclusion】A method of accessing the leg soundness has been proposed by scoring the joint of limb bones in pigs, and it is a crucial supplement method to access the leg soundness. Because toe, leg and gait scores are not correlation with the joint scores, they can’t replace the joint scores. The association analysis results excluded the ENSSSCG00000023160 gene as candidate gene to leg soundness, but both HMGA1 and C6orf106 genes were not excluded. Therefore, the two genes are worthy for further investigations.

The Safety Evaluation of Cefquinome Sulfate Intramammary Infusion (Dry Cow)

WU Tong, ZHANG Dao-kang, LIU Mao-lin, HUANG Hui-li, ZHANG Ning, XU Fei, LIU Yi-ming

Scientia Agricultura Sinica. 2016, 49(20):  4040-4046.  doi:10.3864/j.issn.0578-1752.2016.20.017

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【Objective】The objective of this study is to evaluate the effect of cefquinome sulfate intramammary infusion(dry cow) on the body temperature, milk yield per day, somatic cell counts and mammary microbiota of the health dairy cows.【Method】Six primiparous and multiparous healthy dairy cow were selected, which have not been treated with any antibiotics by systemic or intramammary administration 30 days before. They were fed with food and water without any antibiotics and under normal conditions. Rectal temperature, milk yield per day and somatic cell counts per quarter were recorded on 1 and 0 day before administration, and pathogenic bacteria were isolated and identified. After a single intramammary administration of Cefquinome Sulfate Intramammary Infusion (Dry Cow) to each quarter, milk yield was recorded on 1, 3, 5, 7 and 10 day after administration, rectal temperature per cow and somatic cell counts per quarter were recorded on 12 h, 3, 5, 7 and 10 day after administration, and pathogenic bacteria were isolated and identified on 10 day after administration. The data of rectal temperature, milk yield per day, somatic cell counts and pathogenic bacteria before and after administration were analyzed by SPSS software and the clinical symptom and adverse effects, such as red, swelling, heat and pain were observed continuously throughout the trial. 【Result】The average daily milk yield were 28.8, 27.7, 28.1, 28.7, 28.8, 29.2, and 29.6 kg on 1, 0 day before administration and 1, 3, 5, 7, and 10 day after administration, and it had no significant difference (P>0.05). The somatic cell counts per quarter at different time-points were all in the range of 300-500 thousand per milliliter. There were also no obvious difference in the average rectal temperature of 38.3, 38.4, 38.3, 38.3, 38.3, 38.2 and 38.3℃ at different time-points. The results for bacteria showed that totally 8 strains of E.coli, 5 strains of Streptococcus and 7 strains of Staphylococcus were isolated from the milk samples before administration, but only 1 strain of Streptococcus, 1 strain of Staphylococcus, and 1 strain of E.coli were isolated on 10 day after administration, and no new species of bacteria were found.【Conclusion】It was concluded that cefquinome sulfate intramammary infusion(Dry Cow) is safe for dairy cow, since it has no adverse effect on rectal temperature, milk yield and somatic cell counts.

Appropriate Level of Sodium Selenite for Apis mellifera ligustica Worker Bee Larvae Feed

SU Zhong-qu, ZHANG Wei-xing, CHI Xue-peng, WANG Hong-fang, XU Bao-hua

Scientia Agricultura Sinica. 2016, 49(20):  4047-4055.  doi:10.3864/j.issn.0578-1752.2016.20.018

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【objective】The objective of this study is to find an appropriate level of sodium selenite which added in the food of Apis mellifera ligustica worker larvae, and to provide a theoretical basis for the nutritional needs of Se.【Method】A total of 1 440 one-day old larvae were randomly allotted into 2 batches, each batch involve 720 larvae, one of the batch was used to measure the pupation rate and eclosion rate, the other was used to measure enzyme activity and molecular index. Two batches were all randomly divided into 6 groups, the control group was fed basic diet and the treatment groups were fed different diets with sodium selenite at the levels of 0.2, 0.4, 0.6, 0.8, and 1.0 mg·kg^-1, respectively. Each group had 5 repetitions and involved 24 larvae in each. The 3-, 5- and 7-day-old larvae were used to measure larvae weight, T-AOC (total antioxidant capacity), T-SOD (total superoxide dismutase), MDA (malondialdehyde), and PO (phenoloxidase). The gene expression of SelD, SerRs, Sbp1 and Sbp2 were measured by using 5-day-old larvae. The pupation rate was calculated at 7-day-old.【Result】Compared with the control group, when the added levels of sodium selenite were 0.2-0.6 mg·kg^-1, the T-AOC was significantly increased (P＜0.05), when the added levels of sodium selenite were 0.2-0.8 mg·kg^-1, the activity of T-SOD was significantly increased (P＜0.05) and the MDA content of all the treatment groups was significantly decreased (P＜0.05). When the added level of sodium selenite was 0.4 mg·kg^-1, the activity of PO was significantly increased (P＜0.05). When the added level of sodium selenite was 0.6 mg·kg^-1, the gene expression levels of SelD, SerRs, Sbp1and Sbp2 was significantly higher than the control in 5-day-old larvae (P＜0.05). When the added level of sodium selenite was 0.4 mg·kg^-1, the weight before pupation was significantly higher than the control (P＜0.05). Compared with the control group, when the added level of sodium selenite was 0.2 mg·kg^-1, the pupation rate were significantly increased (P＜0.05). 【Conclusion】Under artificial condition, when the selenium level of basic diet was 0.21 mg·kg^-1, the appropriate level of sodium selenite which added in the food of A. m. ligusticaby the fitting curve which involve the weight before pupation and the pupation rate, and it means that the appropriate level of selenium which added in the food of A. m. ligustica worker larvae is 0.32-0.36 mg·kg^-1. worker larvae was 0.24-0.33 mg·kg^-1

RESEARCH NOTES

Effect of Maize Row Spacing on Biomass, Root Bleeding Sap and Nutrient of Soybean in Relay Strip Intercropping Systems

YANG Feng, LOU Ying, LIU Qin-lin, FAN Yuan-fang, LIU Wei-guo, YONG Tai-wen, WANG Xiao-chun, YANG Wen-yu

Scientia Agricultura Sinica. 2016, 49(20):  4056-4064.  doi:10.3864/j.issn.0578-1752.2016.20.019

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【Objective】 Soybean (Glycine max (L.) Merr.) is an important N-fixing crop, and is a major oilseed crop produced and consumed for protein and oil throughout the world. How maize planting patterns affect the root biomass, bleeding sap and nutrient of soybean were analyzed under maize-soybean relay strip intercropping systems. This study will provide a theoretical basis for reasonable group configuration of maize-soybean strip intercropping. 【Method】The experiment was conducted in 2012-2013 to analyze the effect of maize row-spacing patterns on soybean root biomass, accumulation of N, P and K, bleeding intensity and bleeding sap components at V3, V5 and R2 stages under maize- soybean relay strip intercropping systems. This experiment comprised three maize and soybean intercropping systems and one sole soybean treatment with three replications. Maize cultivar Chuandan418, and soybean cultivar Nandou12 were used as materials. The following maize planting patterns were adopted: A1 (20 cm+180 cm, 20 cm narrow row and 180 cm wide row), A2 (40 cm +160 cm), A3 (60 cm+140 cm). Maize-to-soybean row ratio was also 2:2. Soybean was planted in the wide rows before the reproductive stage of maize. 【Result】 Above-ground biomass, below-ground biomass and root accumulation of N, P and K in soybean were decreased with increasing maize narrow-row spacing, and these parameters under intercropping conditions were less than those of monoculture significantly (P<0.05). Root bleeding intensity of soybean was decreased from V3 to R2 stages, opposite trends were appeared from A1 to A3 treatments at the same growth stage. No significant difference of root bleeding intensity was found in A1 and A2 treatments (P>0.05). Root bleeding intensity at V3, V5 and R2 stage in A1 treatment was decreased by 27.69%, 26.11% and 23.23% compared with those of monoculture, respectively. Nitrate nitrogen, total phosphorus, total potassium, soluble sugar content of bleeding sap and bleeding intensity of soybean root increased gradually with increasing the soybean growth stages except the ammonium nitrogen content. The correlation among root nutrient accumulation, bleeding intensity, bleeding sap component concentration, above-ground and below-ground biomass were significant (P < 0.01). 【Conclusion】Therefore，planting pattern affect the soybean biomass and root bleeding intensity in maize-soybean relay strip intercropping system, this will influence the nutrient absorption and matter transport of soybean root.