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    Crop Genetics · Breeding · Germplasm Resources
    Combining QTL mapping and expression profile analysis to identify candidate genes of cold tolerance from Dongxiang common wild rice (Oryza rufipogon Griff.)
    ZHAO Jie, QIN Jing-jing, SONG Qian, SUN Chuan-qing, LIU Feng-xia
    2016, 15(9): 1933-1943.  DOI: 10.1016/S2095-3119(15)61214-X
    Abstract ( )   PDF in ScienceDirect  
    Rice (Oryza sativa L.), a tropical and subtropical crop, is susceptible to low temperature stress during seedling, booting, and flowering stages, which leads to lower grain quality levels and decreasing rice yields. Cold tolerance is affected by multiple genetic factors in rice, and the complex genetic mechanisms associated with chilling stress tolerance remain unclear. Here, we detected seven quantitative trait loci (QTLs) for cold tolerance at booting stage and identified one cold tolerant line, SIL157, in an introgression line population derived from a cross between the indica variety Guichao 2, as the recipient, and Dongxiang common wild rice, as the donor. When compared with Guichao 2, SIL157 showed a stronger cold tolerance during different growth stages. Through an integrated strategy that combined QTL-mapping with expression profile analysis, six candidate genes, which were up-regulated under chilling stress at the seedling and booting developmental stages, were studied. The results may help in understanding cold tolerance mechanisms and in using beneficial alleles from wild rice to improve the cold tolerance of rice cultivars through molecular marker-assisted selection.
    Phenotypic characterization and fine mapping of mps1, a premature leaf senescence mutant in rice (Oryza sativa L.)
    LIU Zhong-xian, CUI Yu, WANG Zhong-wei, XIE Yuan-hua, SANG Xian-chun, YANG Zheng-lin, ZHANG Chang-wei, ZHAO Fang-ming, HE Guang-hua, LING Ying-hua
    2016, 15(9): 1944-1954.  DOI: 10.1016/S2095-3119(15)61279-5
    Abstract ( )   PDF in ScienceDirect  
       Leaves play a key role in photosynthesis in rice plants. The premature senescence of such plants directly reduces the accumulation of photosynthetic products and also affects yield and grain quality significantly and negatively. A novel premature senescence mutant, mps1 (mid-late stage premature senescence 1), was identified from a mutant library consisting of ethyl methane sulfonate (EMS) induced descendants of Jinhui 10, an elite indica restorer line of rice. The mutant allele, mps1, caused no phenotypic differences from the wild type (WT), Jinhui 10, but drove the leaves to turn yellow when mutant plants grew to the tillering stage, and accelerated leaf senescence from the filling stage to final maturation. We characterized the agronomic traits, content of photosynthetic pigments and photosynthetic efficiency of mps1 and WT, and fine-mapped MPS1. The results showed that the MPS1-drove premature phenotype appeared initially on the leaf tips at the late tillering stage and extended to the middle of leaves during the maturing stage. Compared to the WT, significant differences were observed among traits of the number of grains per panicle (–31.7%) and effective number of grains per panicle (–38.5%) of mps1 individuals. Chlorophyll contents among the first leaf from the top (Top 1st), the second leaf from the top (Top 2nd) and the third leaf from the top (Top 3rd) of mps1 were significantly lower than those of WT (P<0.05), and the levels of photosynthetic efficiency from Top 1st to the forth leaf from the top (Top 4th) of mps1 were significantly lower than those of WT (P<0.01). Results from the genetic analysis indicated that the premature senescence of mps1 is controlled by a recessive nuclear gene, and this locus, MPS1 is located in a 37.4-kb physical interval between the markers Indel145 and Indel149 on chromosome 6. Genomic annotation suggested eight open reading frames (ORFs) within this physical region. All of these results will provide informative references for the further researches involving functional analyses and molecular mechanism exploring of MPS1 in rice.
    Mapping QTLs for stomatal density and size under drought stress in wheat (Triticum aestivum L.)
    WANG Shu-guang, JIA Shou-shan, SUN Dai-zhen, FAN Hua, CHANG Xiao-ping, JING Rui-lian
    2016, 15(9): 1955-1967.  DOI: 10.1016/S2095-3119(15)61264-3
    Abstract ( )   PDF in ScienceDirect  
       Stomatal density and size affect plant water use efficiency, photosynthsis rate and yield. The objective of this study was to gain insights into the variation and genetic basis of stomatal density and size during grain filling under drought stress (DS) and well-watered (WW) conditions. The doubled haploid population derived from a cross of wheat cultivars Hanxuan 10 (H10), a female parent, and Lumai 14 (L14), a male parent, was used for phenotyping at the heading, flowering, and mid- and late grain filling stages along with established amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers. The stomatal density of doubled haploid (DH) lines was gradually increased, while the stomatal lengths and widths were gradually decreased during grain filling stage. Twenty additive QTLs and 19 pairs of epistatic QTLs for the 3 traits were identified under DS. The other 20 QTLs and 25 pairs epistatic QTLs were obtained under WW. Most QTLs made more than 10% contributions to the total phenotypic variations at one growth stage under DS or WW. Furthermore, QTLs for stomatal density near Xwmc74 and Xgwm291 located on chromosome 5A were tightly linked to previously reported QTLs regulating total number of spikelets per spike, number of sterile spikelets per spike and proportion of fertile spikelets per spike. Qsw-2D-1 was detected across stages, and was in the same marker region as a major QTL for plant height, QPH.cgb-2D.1. These indicate that these QTLs on chromosomes 5A and 2D are involved in regulating these agronomic traits and are valuable for molecular breeding.
    A plastidic ATP/ADP transporter gene, IbAATP, increases starch and amylose contents and alters starch structure in transgenic sweetpotato
    WANG Yan-nan, LI Yan, ZHANG Huan, ZHAI Hong, LIU Qing-chang, HE Shao-zhen
    2016, 15(9): 1968-1982.  DOI: 10.1016/S2095-3119(15)61192-3
    Abstract ( )   PDF in ScienceDirect  
        A plastidic adenosine triphosphate (ATP)/adenosine diphosphate (ADP) transporter (AATP) is responsible for importing ATP from the cytosol into plastids. In dicotyledonous plants, increasing ATP supply is a potential way to facilitate anabolic synthesis in heterotrophic plastids. In this study, a gene encoding the AATP protein, named IbAATP, was isolated from sweetpotato (Ipomoea batatas (L.) Lam.). Transcripts of IbAATP were predominantly detected in the storage roots and leaves and were induced by exogenous sucrose and subjected to circadian rhythm. Transient expression of IbAATP in tobacco and onion epidermal cells revealed the plastidic localization of IbAATP. The overexpression of IbAATP in sweetpotato significantly increased the starch and amylose contents and led to enlarged starch granules. The IbAATP-overexpressing plants showed altered fine structure of amylopectin, which contained an increased proportion of chains with a degree of polymerization (DP) of 10–23 and a reduced number of chains with a DP of 5–9 and 24–40. In addition, starch from the transgenic plants exhibited different pasting properties. The transcript levels of starch biosynthetic genes, including IbAGP, IbGBSSI, IbSSI-IV, and IbSBE, were differentially regulated in the transgenic plants. These results revealed the explicit role of IbAATP in the starch biosynthesis of sweetpotato and indicated that this gene has the potential to be used to improve starch content and quality in sweetpotato and other plants.
    Path analysis and estimation of additive and epistatic gene effects of barley SSD lines
    Jan Bocianowski, Katarzyna Górczak, Kamila Nowosad, Wojciech Rybiński, Dariusz Piesik
    2016, 15(9): 1983-1990.  DOI: 10.1016/S2095-3119(15)61243-6
    Abstract ( )   PDF in ScienceDirect  
        In the paper presented, 99 single seed descent barley lines as well as their parental forms Roland and Apex were studied. The grain weight components and their interrelations were analyzed using simple coefficients of correlation. The direct and indirect effects of such components on grain weight per plant and 1 000-grain weight were estimated using path analysis. In the 2006 and 2007, the spike length and number of spikelets per spike were the determinants of grain weight. Genetic parameters as additive and epistasis effects were estimated for all studied traits. The results indicate the importance of both additive and epistasis gene effects of number of spikes per plant, grain weight per spike, grain number per plant and grain weight per plant in this study.
    Genetic diversity of pepper (Capsicum spp.) germplasm resources in China reflects selection for cultivar types and spatial distribution
    ZHANG Xiao-min, ZHANG Zheng-hai, GU Xiao-zhen, MAO Sheng-li, LI Xi-xiang, Jo?l Chadoeuf, Alain Palloix, WANG Li-hao, ZHANG Bao-xi
    2016, 15(9): 1991-2001.  DOI: 10.1016/S2095-3119(16)61364-3
    Abstract ( )   PDF in ScienceDirect  
        Pepper (Capsicum spp.) is an important vegetable crop in the world. Now the pepper in China contributes one-third of the world’s peppers production. Genetic diversity of the pepper germplasm of China is expected interesting to know. To explore the structure of genetic diversity in Chinese pepper germplasm resources and possible relationship with cultivar types or geographic origin, we sampled and compared 372 GenBank pepper accessions (local cultivars and landraces) from 31 provinces, autonomous regions and municipalities of China and 31 additional accessions from other countries. These accessions were genotyped using 28 simple sequence repeat (SSR) markers spanning the entire pepper genome. We then investigated the genetic structure of the sampled collection using model-based analysis in STRUCTURE v2.3.4 and examined genetic relationships by the unweighted pair-group method of mathematical averages (UPGMA) in MEGA. In addition to geographic origin, we evaluated eight plant and fruit traits. In total, 363 alleles were amplified using the 28 SSR primers. Gene diversity, polymorphism information content and heterozygosity of the 28 SSR loci were estimated as 0.09–0.92, 0.08–0.92 and 0.01–0.34, respectively. The UPGMA cluster analysis clearly distinguished Capsicum annuum L. from other cultivated pepper species. Population structure analysis of the 368 C. annuum accessions uncovered three genetic groups which also corresponded to distinct cultivar types with respect to the plant and fruit descriptors. The genetic structure was also related to the geographic origin of the landraces. Overall results indicate that genetic diversity of Chinese pepper landraces were structured by migration of genotypes followed by human selection for cultivar types in agreement with consumption modes and adaptation to the highly diversified agro-climatic conditions.
    Varietal difference in the correlation between leaf nitrogen content and photosynthesis in rice (Oryza sativa L.) plants is related to specific leaf weight
    LIU Xi, LI Yong
    2016, 15(9): 2002-2011.  DOI: 10.1016/S2095-3119(15)61262-X
    Abstract ( )   PDF in ScienceDirect  
        Increasing leaf photosynthesis per area (A) is of great importance to achieve yield further improvement. The aim of this study was to exploit varietal difference in A and its correlation with specific leaf weight (SLW). Twelve rice cultivars, including 6 indica and 6 japonica varieties, were pot-grown under two N treatments, low N (LN) and sufficient N (SN). Leaf photosynthesis and related parameters were measured at tillering stage. Compared with LN treatment, A, stomatal conductance (gs), mesophyll conductance (gm), leaf N content (Narea), and chlorophyll content were significantly improved under SN treatment, while SLW and photosynthetic N use efficiency (PNUE) were generally decreased. Varietal difference in A was positively related to both gs and gm, but not related to Narea. This resulted in a low PNUE in high Narea leaves. Varietal difference in PNUE was generally negatively related to SLW. Response of PNUE to N supply varied among different rice cultivars, and interestingly, the decrease in PNUE under SN was negatively related to the decrease in SLW. With a higher Narea, japonica rice cultivars did not show a higher A than indica rice cultivars because of possession of high-SLW leaves. Therefore, varietal difference in A was not related to Narea, and SLW can substantially interfere with the correlation between A and Narea. These findings may provide useful information for rice breeders to maximize A and PNUE, rather than over reliance on Narea as an indicator of photosynthetic performance.
    Effects of CO2 enrichment and spikelet removal on rice quality under open-air field conditions
    JING Li-quan, WU Yan-zhen, ZHUANG Shi-teng, WANG Yun-xia, ZHU Jian-guo, WANG Yu-long, YANG Lian-xin
    2016, 15(9): 2012-2022.  DOI: 10.1016/S2095-3119(15)61245-X
    Abstract ( )   PDF in ScienceDirect  
        The increase of atmospheric carbon dioxide (CO2) concentration adversely affect several quality traits of rice grains, but the biochemical mechanism remains unclear. The objectives of this study were to determine how changes in the source-sink relationship affected rice quality. Source-sink manipulation was achieved by free-air CO2 enrichment from tillering to maturity and partial removal of spikelet at anthesis using a japonica rice cultivar Wuyunjing 23. Enrichment with CO2 decreased the head rice percentage and protein concentration of milled rice, but increased the grain chalkiness. In contrast, spikelet removal resulted in a dramatic increase in the head rice percentage and protein concentration, and much less grain chalkiness. Neither CO2 enrichment nor spikelet removal affected the starch content, but the distribution of starch granule size showed distinct treatment effects. On average, spikelet removal decreased the percentage of starch granules of diameter >10 and 5–10 μm by 23.6 and 5.6%, respectively, and increased those with a diameter of 2–5 and <2 μm by 4.6 and 3.3%, respectively. In contrast, CO2 elevation showed an opposite response: increasing the proportion of large starch granules (>5 μm) and decreasing that of <5 μm. The starch pasting properties were affected by spikelet removal much more than by CO2 elevation. These results indicated that the protein concentration and starch granule size played a role in chalkiness formation under these experimental conditions.
    Flavonoid content and expression analysis of flavonoid biosynthetic genes in herbaceous peony (Paeonia lactiflora Pall.) with double colors
    WU Yan-qing, WEI Meng-ran, ZHAO Da-qiu, TAO Jun
    2016, 15(9): 2023-2031.  DOI: 10.1016/S2095-3119(15)61318-1
    Abstract ( )   PDF in ScienceDirect  
        Herbaceous peony (Paeonia lactiflora Pall.) is a famous flower with medicinal values, and its flowers have a number of medicinal constituents, especially flavonoids. In this study, a P. lactiflora cultivar with double colors including white outer-petal and yellow inner-petal was used as the experimental materials to perform the qualitative and quantitative analysis of flavonoids by high-performance liquid chromatograph-electrospray ionization-mass spectrometry (HPLC-ESI-MSn) and investigate the expression patterns of flavonoid biosynthetic genes using real-time quantitative polymerase chain reaction (Q-PCR). The results showed that the colors of both petals gradually weakened with flower development. Moreover, one main anthocyanin composition (peonidin 3,5-di-O-glucoside) and five main anthoxanthin compositions (kaempferol di-hexoside, kaempferol-3-O-malonylglucoside-7-O-glucoside, quercetin-3-O-galactoside, luteolin-7-O-glucoside and isorhamnetin-3-O-glucoside) were found in the both, differing significantly in their peak areas only. Total anthocyanin, anthoxanthin and flavonoid contents in white outer-petal and yellow inner-petal gradually decreased during flower development, and were consistently higher in white outer-petal. Furthermore, the expression patterns of nine structural genes in P. lactiflora flavonoid biosynthetic pathway showed that the expression levels of phenylalanine ammonialyase gene (PlPAL), chalcone synthase gene (PlCHS), flavanone 3-hydroxylase gene (PlF3H), anthocyanidin synthase gene (PlANS) and UDP-glucoside: flavonoid 5-O-glucosyltransferase gene (PlF5GT) in two petals basically presented declined tendencies, and transcription levels of PlPAL, PlCHS, PlANS, PlF3GT and PlF5GT also tended to be higher in white outer-petal, which was correlated with their flavonoid contents. These results would lay a solid foundation for the exploration and utilization of flavonoid resources in P. lactiflora flowers.
    Plant Protection
    The sigma 54 genes rpoN1 and rpoN2 of Xanthomonas citri subsp. citri play different roles in virulence, nutrient utilization and cell motility
    Gibson Kamau Gicharu, SUN Dong-ling, HU Xun, FAN Xiao-jing, ZHUO Tao, WU Chuan-wan, ZOU Hua-song
    2016, 15(9): 2032-2039.  DOI: 10.1016/S2095-3119(15)61317-X
    Abstract ( )   PDF in ScienceDirect  
        The sigma factor 54 (σ54) controls the expression of many genes in response to nutritional and environmental conditions. There are two σ54 genes, rpoN1 (XAC1969) and rpoN2 (XAC2972), in Xanthomonas citri subsp. citri. To investigate their functions, the deletion mutants ΔrpoN1, ΔrpoN2 and ΔrpoN1N2 were constructed in this study. All the mutants delayed canker development in low concentration inoculation in citrus plants. The bacterial growth of mutants was retarded in the medium supplemented with nitrogen and carbon resources. Under either condition, the influence degree caused by deletion of rpoN2 was larger than the deletion of rpoN1. Remarkably, the mutant ΔrpoN1 showed a reduction in cell motility, while the mutant ΔrpoN2 increased cell motility. Our data suggested that the rpoN1 and rpoN2 play diverse roles in X. citri subsp. citri.
    A novel strategy to enhance resistance to Cucumber mosaic virus in tomato by grafting to transgenic rootstocks
    BAI Miao, CHEN Wen-ting, XIE Bing-yan, YANG Guo-shun
    2016, 15(9): 2040-2048.  DOI: 10.1016/S2095-3119(16)61330-8
    Abstract ( )   PDF in ScienceDirect  
       Cucumber mosaic virus (CMV) can infect a wide range of host species. For the lacking of CMV resistant varieties of tomato, RNA interference (RNAi) can be used as a fast and effective method for the generation of transgenic resistant varieties. In this current study, five intron-spliced hairpin RNA (ihpRNA) plant expression vectors aimed at five genes of CMV have been constructed. Transgenic tomatoes were obtained by Agrobacterium tumefaciens-mediated transformation with expression vectors. Highly resistant generations of transgenic plants were employed as rootstocks and grafted onto non-transgenic tomatoes that resulted in the successful transfer of resistance to the scions. Using a novel method of plant cuttings for rootstock propagation, we obtained large quantities of disease-resistant material. Further, this method produces scions that can remain undetectable for transgenic resistance marker genes that may provide novel approaches to evade collective concerns about genetically-modified organism (GMO) biosafety.
    Identification of candidate soil microbes responsible for small-scale heterogeneity in strawberry plant vigour
    WEI Feng, FAN Rong, Thomas Passey, HU Xiao-ping, Xiangming Xu
    2016, 15(9): 2049-2058.  DOI: 10.1016/S2095-3119(16)61354-0
    Abstract ( )   PDF in ScienceDirect  
        Studies were conducted to identify candidate soil microbes responsible for observed differences in strawberry vigour at a small spatial scale, which was not associated with visual disease symptoms. Samples were obtained from the soils close to the rhizosphere of ‘big’ and ‘small’ plants from small plots which exhibited large local heterogeneity in plant vigour. A metabarcoding approach was used to profile bacterial and fungal compositions, using two primer pairs for 16S ribosomal RNA genes (16S rDNA) and one for the fungal internal transcribed spacer (ITS) region. Of the two 16S rDNA primer sets, the 341F/805R resulted in sequences of better quality. A total 28 operational taxonomic units (OTUs) had differential relative abundance between samples from ‘big’ and ‘small’ plants. However, plausible biological explanation was only possible for three fungal OTUs. Two were possible phytopathogens: Verticillium spp. and Alternaria alternata although the latter has never been considered as a main pathogen of strawberry in the UK. For samples from ‘small’ plants, the abundance of these OTUs was much greater than from ‘big’ plants. The opposite was true for a mycorrhizal OTU. These results suggest that soil microbes related to crop production can be identified using metabarcoding technique. Further research is needed to assess whether A. alternata and Verticillium spp. could affect strawberry growth in the field.
    Thrips-mediated impacts from transgenic rice expressing Cry1Ab on ecological fitness of non-target predator Orius tantilus (Hemiptera: Anthocoridae)
    Akhtar Zunnu Raen, DANG Cong, WANG Fang, PENG Yu-fa, YE Gong-yin
    2016, 15(9): 2059-2069.  DOI: 10.1016/S2095-3119(16)61414-4
    Abstract ( )   PDF in ScienceDirect  
        Various rice lines have been genetically modified with genes from Bacillus thuringiensis (Bt) to continuously produce Bt insecticidal proteins against lepidopteran larvae. The Bt insecticidal protein constantly expresses in the plants to create an opportunity for non-target herbivores to acquire and convey the protein to their predators or parasitoids across trophic levels. This paper evaluates the effects of Bt rice (namely, Kemindao 1 (KMD1) and Kemindao 2 (KMD2)) expressing Cry1Ab as compared to its non-Bt control line, Xiushui 11 on non-target predator Orius tantilus (a generalist predatory anthocorid of thrips) under laboratory and field conditions. To measure several biological parameters such as total nymphal duration and fecundity of this bug, it was reared on thrips and pollens of KMD1 and KMD2 as compared to their control under laboratory conditions. By comparison with the control, Bt rice did not significantly affect main life-history characteristics (total nymphal duration, female adult longevity, oviposition period and fecundity) of this anthocorid preying on Bt rice-fed thrips along with Bt rice pollens, except that the fecundity of this predator for KMD1 was distinctly lower as compared with KMD2 or the control. Enzyme-linked immunosorbent assay (ELISA) results showed that no Cry1Ab protein was detected in this predator fed on thrips or rice pollen from Bt rice but was in Bt rice pollens. With the beat plate, plastic bag and color trap sampling methods, two-year field monitoring of O. tantilus abundance demonstrated that Bt rice had no significant detrimental effects on the population dynamics and seasonal average densities of this predatory anthocorid as compared with the control. Thus, it is suggested that growing our tested Bt rice (KMD1 and KMD2) producing Cry1Ab will pose a negligible risk to the anthocorid, O. tantilus.
    Animal Science · Veterinary Science
    An efficient and rapid method to detect and verify natural antisense transcripts of animal genes
    Zhang Li, Zhao Rui, Xiao Mei, Lin Shu-dai, Li Bi-xiao, Qiu Feng-fang, Ma Jing-e, Zhang Dexiang, Nie Qing-hua, An Li-long, Zhang Xi-quan
    2016, 15(9): 2070-2076.  DOI: 10.1016/S2095-3119(15)61266-7
    Abstract ( )   PDF in ScienceDirect  
        High-throughput sequencing has identified a large number of sense-antisense transcriptional pairs, which indicates that these genes were transcribed from both directions. Recent reports have demonstrated that many antisense RNAs, especially lncRNA (long non-coding RNA), can interact with the sense RNA by forming an RNA duplex. Many methods, such as RNA-sequencing, Northern blotting, RNase protection assays and strand-specific PCR, can be used to detect the antisense transcript and gene transcriptional orientation. However, the applications of these methods have been constrained, to some extent, because of the high cost, difficult operation or inaccuracy, especially regarding the analysis of substantial amounts of data. Thus, we developed an easy method to detect and validate these complicated RNAs. We primarily took advantage of the strand specificity of RT-PCR and the single-strand specificity of S1 endonuclease to analyze sense and antisense transcripts. Four known genes, including mouse β-actin and Tsix (Xist antisense RNA), chicken LXN (latexin) and GFM1 (G elongation factor, mitochondrial 1), were used to establish the method. These four genes were well studied and transcribed from positive strand, negative strand or both strands of DNA, respectively, which represented all possible cases. The results indicated that the method can easily distinguish sense, antisense and sense-antisense transcriptional pairs. In addition, it can be used to verify the results of high-throughput sequencing, as well as to analyze the regulatory mechanisms between RNAs. This method can improve the accuracy of detection and can be mainly used in analyzing single gene and was low cost.
    Effects of soybean meal replacement with fermented alfalfa meal on the growth performance, serum antioxidant functions, digestive enzyme activities, and cecal microflora of geese
    YIN Hai-cheng, HUANG Jin
    2016, 15(9): 2077-2086.  DOI: 10.1016/S2095-3119(15)61198-4
    Abstract ( )   PDF in ScienceDirect  
        Fermented forages are important feedstuffs. Bacillus subtilis inoculants are often used to improve the value of forage legume fermentation. The present work was conducted to study the effects of replacing soybean meal with solid-state fermented alfalfa meal (FAM) with B. subtilis ACCC 01746 on growth performance, serum antioxidant and digestive enzyme activities, and cecal microflora in goose. 300 healthy geese with similar body weights were randomly assigned to six treatment groups with five replicates of 10 geese (five males and five females) each. Geese were fed ad libitum for 35 days. Results showed that the geese fed with 4 and 8% FAM exhibited no significant effects on their final body weights (FBW) and average day gain (ADG) (P>0.05), whereas 12% or higher FAM caused poor growth of the geese compared with control diet (linear (L): P<0.05). The average daily feed intake (ADFI) (quadratic (Q): P<0.05) and feed conversion ratio (FCR) (L: P<0.05) with 8% or higher supplementation level were higher than those of the control group. The activities of antioxidant enzyme in serum increased, and the level of malondialdehyde (MDA) significantly decreased with increasing dietary FAM level (L: P<0.05). However, no significant differences were observed at 8% or lower supplementation level for glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) (P>0.05) and at 4% for catalase (CAT) supplementation level compared with the control group. All diets containing FAM increased digestive enzyme activities in geese. However, geese fed diets with 12% FAM supplementation showed the highest trypsin activities in pancreas (Q: P<0.05). Supplementation with 12% or higher FAM significantly increased amylase activities in pancreas (L: P<0.05) and duodenum (L: P<0.05) compared with the control group. Significant differences were not observed in total anaerobic bacteria between geese fed with FAM and control diets on day 35 (P>0.05). The numbers of Bifidobacterium and Lactobacillus in the cecum of geese fed with FAM significantly increased (L: P<0.05), but no significant effects were found with 4 and 8% FAM supplementation levels compared with the control (P>0.05). By contrast, the coliform counts of cecum decreased with increasing inclusion of FAM, but these counts were significantly reduced in geese fed diets with 12% or higher FAM supplementation level (L: P<0.05). Collectively, our results indicated that supplementation of the basal geese diet with 8% FAM had no apparent adverse effects on growth performance, serum antioxidant enzyme activities, and digestive parameters and beneficial microbiota.
    Effects of Lactobacillus plantarum and fibrolytic enzyme on the fermentation quality and in vitro digestibility of total mixed rations silage including rape straw
    LIU Qin-hua, LI Xiang-yu, Seare T Desta, ZHANG Jian-guo, SHAO Tao
    2016, 15(9): 2087-2096.  DOI: 10.1016/S2095-3119(15)61233-3
    Abstract ( )   PDF in ScienceDirect  
       The aim of the current study was to investigate the effects of a lactic acid bacteria inoculant (Lactobacillus plantarum, LP), fibrolytic enzyme (EN), combination of LP and EN (LP+EN) on fermentation quality, nutritive characteristics and in vitro digestibility of total mixed ration (TMR) silages containing 0, 7.5 and 15.0% (on dry matter basis) of rape (Brassica campestris L.) straw (RS) (denoted as CTMR, LTMR and MTMR, respectively).  After ensiling for 60 days, TMR silages without additives were well preserved, but MTMR had higher pH than CTMR and LTMR. There were no differences in other parameters of fermentation quality, microbial composition, nutrition and in vitro digestibility between CTMR and LTMR except for yeast and mold number and crude protein (CP) content. CTMR and LTMR silage had higher CP content, in vitro neutral detergent fiber digestibility (IVNDFD) and in vitro acid detergent fiber digestibility (IVADFD), lower acid detergent fiber (ADF) content than MTMR silage.  LP and EN decreased pH and increased dry matter (DM) recovery of TMR silages. LP+EN improved the fermentation quality, nutritive characteristics and in vitro digestibility of TMR silages, showed by lower pH, ADF content, higher lactic acid content, in vitro dry matter digestibility (IVDMD) and IVNDFD.  Therefore, It was suggested that TMR silage contained 7.5% RS on a DM basis and treated with LP+EN can be as a useful feed for ruminant.
    Reference genes for quantitative real-time PCR analysis and quantitative expression of P5CS in Agropyron mongolicum under drought stress
    TIAN Qing-song, WANG Shu-yan, DU Jian-cai, WU Zhi-juan, LI Xiao-quan, HAN Bing
    2016, 15(9): 2097-2104.  DOI: 10.1016/S2095-3119(15)61238-2
    Abstract ( )   PDF in ScienceDirect  
        Reference genes, stably expressing in different tissues and cells, are commonly used as the references in expression analysis. Selecting the optimum reference gene is crucial to the success of experiments. In this study, the expression stabilities of nine common reference genes, including ACT2, 18S rRNA, APRT, EF-1α, RNA POL II, TUBα, TUBβ, GAPDH and TLF of Agropyron mongolicum, were studied under drought condition. Among them, 18S rRNA was found to be the most optimum reference gene under drought stress by the analyzing of geNorm and NormFinder software. Quantitative expression levels of P5CS using 18S rRNA as the reference gene, and proline contents under drought stress in A. mongolicum were further operated, and we found the expression level of P5CS gene and proline content had a significantly positive relationship (R2=0.7763, P<0.05). This study established and validated 18S rRNA as the reference genes in A. mongolicum under drought stress, providing a powerful tool for the quantitative expression analysis of drought genes in A. mongolicum.
    A duplex RT-PCR assay for detection of H9 subtype avian influenza viruses and infectious bronchitis viruses
    WEI Yan-di, GAO Wei-hua, SUN Hong-lei, YU Chen-fang, PEI Xing-yao, Sun Yi-peng, LIU Jin-hua, PU Juan
    2016, 15(9): 2105-2113.  DOI: 10.1016/S2095-3119(15)61316-8
    Abstract ( )   PDF in ScienceDirect  
    H9 subtype avian influenza virus (AIV) and infectious bronchitis virus (IBV) are major pathogens circulating in poultry and have resulted in great economic losses due to respiratory disease and reduced egg production. As similar symptoms are elicited by the two pathogens, it is difficult for their differential diagnosis. So far, no reverse transcription-polymerase chain reaction (RT-PCR) assay has been found to differentiate between H9 AIV and IBV in one reaction. Therefore, developing a sensitive and specific method is of importance to simultaneously detect and differentiate H9 AIV and IBV. In this study, a duplex RT-PCR (dRT-PCR) was established. Two primer sets target the hemagglutinin (HA) gene of H9 AIV and the nucleocapsid (N) gene of IBV, respectively. Specific PCR products were obtained from all tested H9 AIVs and IBVs belonging to the major clades circulating in China, but not from AIVs of other subtypes or other in·fectious avian viruses. The sensitivity of the dRT-PCR assay corresponding to H9 AIV, IBV and mixture of H9 AIV and IBV were at a concentration of 1×101, 1.5×101 and 1.5×101 50% egg infective doses (EID50) mL–1, respectively. The concordance rates between the dRT-PCR and virus isolation were 99.1 and 98.2%, respectively, for detection of samples from H9N2 AIV or IBV infected chickens, while the concordance rate was 99.1% for detection of samples from H9N2 AIV and IBV co-infected chickens. Thus, the dRT-PCR assay reported herein is specific and sensitive, and suitable for the differential diagnosis of clinical infections and surveillance of H9 AIVs and IBVs.
    Soil & Fertilization﹒Irrigation﹒Plant Nutrition﹒ Agro-Ecology & Environment
    Subcellular Cd accumulation characteristic in root cell wall of rice cultivars with different sensitivities to Cd stress in soil
    LIU Bin, CHEN Li, CHEN Shi-bao, LI Ning, ZHENG Han, JIN Ke, PANG Huan-cheng, MA Yi-bing
    2016, 15(9): 2114-2122.  DOI: 10.1016/S2095-3119(15)61227-8
    Abstract ( )   PDF in ScienceDirect  
       The variations of grain cadmiun (Cd) concentrations, translocation factors (TFs) of Cd from roots to shoots/grains of six rice cultivars, characterized with different Cd-sensitivities in polluted soil were studied, the selected rice cultivars were Xiangzao 17 (R1), Jiayu 211 (R2), Xiangzao 42 (R3), Zhuliangyou 312 (R4), Zhuliangyou 611 (R5), and Jinyou 463 (R6), respectively. The Cd subcellular distribution and Cd binding characteristics on subcellular fractions of rice root cell wall (CW) were further investigated. The results showed that the rice grain Cd contents varied significantly, with a maximum variation of 47.0% among the cultivars, the largest grain Cd content was observed with cultivar R1 (Cd-sensitivity cultivar) and the smallest with R5 (Cd-tolerance cultivar). The translocation factors of Cd from roots to shoots (TFshoot) and roots to grains (TFgrain) varied greatly among the cultivars. In general, the TFgrain of the cultivars followed the order of R1>R2>R3>R4> R6-R5. The Cd concentration (mg kg–1 FW) in the fraction of root CW, the fraction of cell wall removing pectin (CW-P) and the fraction of cell wall removing pectin and hemicellulose (CW-P-HC) of the cultivars generally followed the order of CW-P>CW>CW-P-HC; the ratios of Cd concentration (mg kg–1 FW) in the fraction of CW-P to that of CW were mostly more than 1.10, while the ratios of Cd concentration in the fraction of CW-P-HC to that of CW were mostly less than 0.60, indicating that Cd was mainly stored in the hemicellulose of the root CW. The ratios of Cd of CW-P-HC to CW generally followed the descending order of R1~R2>R3>R4>R5~R6 for the cultivars, which implied that hemicellulose is probably the main subcellular pool for transferring Cd into rice grain, and it restrains the translocation of Cd from shoot to the grain, especially for the Cd-tolerance cultivars (R5 and R6), the compartmentation of more Cd in hemicellulose in root CW is probably one of the main mechanisms for Cd tolerance of rice cultivars.
    Effect of different water application intensity and irrigation amount treatments of microirrigation on soil-leaching coastal saline soils of North China
    CHU Lin-lin, KANG Yao-hu, WAN Shu-qin
    2016, 15(9): 2123-2131.  DOI: 10.1016/S2095-3119(15)61263-1
    Abstract ( )   PDF in ScienceDirect  
       In coastal regions, Bohai Gulf is one of the most affected areas by salinization. To study the effects of mocrosprinkler irrigation on the characteristics of highly saline sandy loam soil (ECe (saturated paste extract)=22.3 dS m–1; SAR (sodium adsorption ratio)=49.0) of North China, a laboratory experiment was conducted. Five water application intensity (WAI) treatments (1.7, 3.1, 5.3, 8.8, and 10.1 mm h–1), five irrigation amount (IA) treatments (148, 168, 184, 201, and 223 mm) and three time periods of water redistribution (0, 24 and 48 h) were employed in the study. A compounding microsprinkler system was used for the WAI treatments, and a single microsprinkler was used for the IA treatments. The results indicated that, as soil depth increased, soil water content (θ) increased and then slightly decreased; with WAI and IA consistently increasing, the relatively moist region expanded and the average θ increased. Meanwhile, soil ECe increased as soil depth increased, and the zone with low soil salinity expanded as WAI and IA increased. Although the reduction of the average SAR was smaller than that of the average electrical conductivity of the ECe, these variables decreased in similar fashion as WAI and IA increased under microsprinkler irrigation. The average pH decreased as soil depth increased. Longer time periods of water redistribution led to lower salinity and slight expansion of the SAR zone. Considering the effects of leached salts in coastal saline soils, greater WAI and IA values are more advantageous under unsaturated flow conditions, as they cause better water movement in the soil. After leaching due to microsprinkler irrigation, highly saline soil gradually changes to moderately saline soil. The results provide theoretical and technological guidance for the salt leaching and landscaping of highly saline coastal environments.
    A modified vegetation water supply index (MVWSI) and its application in drought monitoring over Sichuan and Chongqing, China
    WU Men-xin, LU Hou-quan
    2016, 15(9): 2132-2141.  DOI: 10.1016/S2095-3119(15)61257-6
    Abstract ( )   PDF in ScienceDirect  
    Based on the principle of vegetation water supply index (VWSI), relative normalized difference vegetation index (RNDVI) and relative land surface temperature (RLST) are proposed, with which a modified vegetation water supply index (MVWSI) is constructed, i.e., . Compared with VWSI, MVWSI eliminates the difference caused by the spatial and temporal distribution of VWSI. MVWSI was applied to monitor drought over Sichuan and Chongqing, China in 2006, suggesting that MVWSI can demonstrate the whole process of drought from its emergence and development to its weakening and disappearance. The relative precipitation index (RPI) is used to examine the effectiveness of MVWSI. The examination result shows that MVWSI and RPI have high consistency in spatial distribution and in-site time series analysis. Compared with vegetation condition index (VCI), temperature condition index (TCI), vegetation temperature condition index (VTCI) and temperature vegetation drought index (TVDI), MVWSI is simpler in calculation and more stable hence more easily realized in application.
    Transcriptome response of wheat Norin 10 to long-term elevated CO2 under high yield field condition
    LIN Yue-bing, SHEN Cheng-guo, LIN Er-da, HAO Xing-yu, HAN Xue
    2016, 15(9): 2142-2152.  DOI: 10.1016/S1671-2927(00)10702
    Abstract ( )   PDF in ScienceDirect  
    The increasing atmospheric carbon dioxide concentration, caused by fossil fuel combustion and deforestation, plays an important role in plant growth and development. Wheat, as a major staple crop, adapts to climate change by tuning its inherent molecular mechanism, which is not well understood. The present study employed the RNA-Seq method to generate transcriptome profiles of the wheat Norin 10 in response to elevated CO2 in comparison with ambient CO2. The 10 895 787 high-quality clean reads of Norin 10 were assembled de novo using Trinity (without a reference genome) resulting in a total of 18 206 candidate transcripts with significant BLAST matches. GO enrichment analysis of Norin 10 at different CO2 concentrations showed that some functional genes related to plastids, precursor metabolites, and energy, thylakoid and photosynthesis were apparently enriched at elevated CO2 (550 μmol mol–1) in contrast to that at ambient CO2 (400 μmol mol–1); these findings were further confirmed by RT-PCR analysis. The findings demonstrated the specific effects of elevated CO2 during long-term period in free air CO2 enrichment (FACE) on transcriptome response of the high yielding wheat variety, Norin 10, which has a large spike.
    The diversity and potential function of endophytic bacteria isolated from Kobreasia capillifolia at alpine grasslands on the Tibetan Plateau, China
    WANG Ying, YANG Cheng-de, YAO Yu-ling, WANG Yu-qin, ZHANG Zhen-fen, XUE Li
    2016, 15(9): 2153-2162.  DOI: 10.1016/S2095-3119(15)61248-5
    Abstract ( )   PDF in ScienceDirect  
    A total of 50 endophytic bacterial isolates were obtained from Kobreasia capillifolia at alpine grasslands in the Eastern Qilian Mountains on the Tibetan Plateau in China.  Based on the sequencing and phylogenetic analysis of 16S rDNA genes, all isolates phylogenetically related closely to Bacillus, Acinetobacter, Stenotrophomonas, Brevundimonas, Arthrobacter, Curtobacterium, Paenibacillus, Plantibacter, Promicromonospora, Serratia, and Microbacterium, among which Bacillus was the predominant genus (47.8% of the total number of endophytic isolates).  These isolates possessed different biological functions.  In 50 endophytic bacteria, 42 isolates produced indole acetic acid (IAA) on King medium.  There were seven isolates showing potency of mineral phosphate solubilization in Pikovaskaia’s (PKO) liquid medium.  Seven isolates exhibited antagonistic effect against Fusarium avenaceum, Colletotrichum coccodes and Phoma foveata.  This was the first report on diversity and plant growth promotion of endophytic bacteria from K. capillifolia on alpine grassland in the Eastern Qilian Mountains, Chain.  It is essential for revealing the relationship among plant, microorganism, and the special environment because the potential function of endophytic bacteria made a contribution to the interactions of plants and endophytic bacteria.
    Food Science
    Analysis of 13 kinds of steroid hormones in raw milk using modified QuEChERS method combined with UPLC-QTOF-MS
    TAN Xin-tong, LI Zeng-mei, DENG Li-gang, ZHAO Shan-cang, WANG Ming-lin
    2016, 15(9): 2163-2174.  DOI: 10.1016/S2095-3119(16)61386-2
    Abstract ( )   PDF in ScienceDirect  
        Thirteen kinds of steroid hormones in raw milk (cow, goat and buffalo milk) were analyzed with ultra performance liquid chromatography-quadrupole time of flight mass spectrometry (UPLC-QTOF-MS) after extraction and cleanup with the modified QuEChERS method. These steroid hormones included 17β-estradiol, estriol, estrone, diethylstilbestrol, progesterone, melengestrol acetate, megestrol acetate, chlormadinone acetate, 19-nortestosterone, metandienone, boldenone, epitestosterone, and testosterone. The limits of detection for the raw milk basing on 3 times the signal to noise ratios (S/N=3) was in range of 0.07-0.51 µg kg–1, and the limits of quantification (basing on S/N=10 method) covered the ranges from 0.23 to 1.7 µg kg–1. With matrix external standard method, the substances presented recoveries over the range 74.2–99.7%. Qualitative analysis was also done in the mass/mass spectrum (MS/MS) mode and each debris structure of 13 kinds of steroid hormones was achieved. The methodology was then applied in real raw milk samples which were collected in several areas of China and the progesterone was detected with high level.
    Quantification and analysis of anthocyanin and flavonoids compositions, and antioxidant activities in onions with three different colors
    ZHANG Shi-lin, DENG Peng, XU Yu-chao, Lü Shan-wu, WANG Jian-jun
    2016, 15(9): 2175-2181.  DOI: 10.1016/S2095-3119(16)61385-0
    Abstract ( )   PDF in ScienceDirect  
        Onion (Allium cepa L.) is a very important vegetable crop widely cultivated throughout the world. In this study, we measured total anthocyanins and flavonols of onions of three colors. The fresh weight (FW) of total anthocyanins were (29.99±1.19), (9.64±1.30) and (0.75±0.40) mg 100 g–1 fruit in red, yellow and white onions, respectively. Likewise, the FW of total flavonoids were (111.10±5.98), (36.64±3.59), and 0 mg 100 g–1 in red, yellow and white, respectively. Four types of anthocyanins (delphinidin 3,5-diglycosides, cyanidin 3,5-diglycosides, cyanidin 3-glycosides and cyanidin 3-(6´´-malonyl)-glucopyranoside) and two kinds of flavonoids (quercetin and quercetin 3-glycosides) were identified in two varieties (red and yellow bulb) of onions by HPLC/DAD-ESI/MS. The total polyphenol contents were also measured by means of Folin-Ciocalteu method. Moreover, all the concentrations of anthocyanins, flavonoids and polyphenols showed significantly positive correlations with antioxidant activities measured by DPPH˙, ABTS˙+ and FRAP assays. This study provided information on anthocyanin and flavonoids compositions that will be useful for onion breeding.