Table of Content

01 April 2014, Volume 47 Issue 7

Previous Issue    Next Issue

CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS

Identification of Indica-Japonica Attribute and Prediction of Heterosis of Zheyou Hybrids Rice Using InDel Molecular Markers

WANG Lin-You, ZHANG Li-Xia, GOU Xiao-Xia, FAN Hong-Huan, JIN Qing-Sheng, WANG Jian-Jun

Scientia Agricultura Sinica. 2014, 47(7):  1243-1255.  doi:10.3864/j.issn.0578-1752.2014.07.001

Abstract ( 533 )   HTML ( 8 )   PDF (682KB) ( 830 )   Save

References | Related Articles | Metrics

【Objective】 It is of great significance to identify indica or japonica accurately and predict the heterosis by genetic distance for super high-yielding rice breeding and exploring the indica-japonica inter-subspecies heterosis of rice. 【Method】The indica-japonica attribute of twelve Zheyou hybrid rice lines and their parents were identified by 19 pairs of InDel molecular markers designed on the basis of the comparative genomic DNA sequences between indica variety 9311 and japonica variety Nipponbare. According to the gene frequency of indica/japonica genomics detected by InDel molecular markers, a method was set up modified upon Lu’s report, and all the tested materials were judged for their different attributes, i.e indica, indica-cline, intermediate indica cline, intermediate japonica-cline, japonica-cline and japonica six types. Genetic distances among 13 rice parents (one sterile line and 12 restorer lines) were assessed by InDel molecular markers and calculated by using Nei’s way. The genetic similarity (GS) was reckoned by using UPGMA method to analyze the cluster of genetic similarity. Principal component analyses (PCA) were conducted using SPSS 17.0 statistical analysis software to InDel band assignments. Then, on the basis of the average values of the eigenvectors of the first and second principal components, a scatterplot was created. The control heterosis of six traits, grain yield, effective panicles per hole, spikelet number per panicle, seed setting rate, 1000-grain weight and grain weight per panicle in hybrid F1 were investigated. Furthermore, the correlations between InDel genetic distance and heterosis related to yield traits were analyzed, and possible relation to the performance of Zheyou hybrid rice lines were discussed. 【Result】 The indica-japonica attributes of the tested materials were identified and the results showed that the sterile line Zhe 04A is identified into japonica type. Eight restoring lines were thought of belonging to indica or indica-cline type, and eight combinations derived from them were judged into a range from intermediate indica-cline to japonica-cline type, it was also verified that these eight combinations were typical indica-japonica combinations. Another four restoring lines were divided into the type of japonica-cline, the four combinations from these lines were judged into japonica type. The cluster analyses were indicated that thirty-five materials could be divided into two major groups, indica group, and japonica group, at 0.350 score of GS. At 0.638 score of GS, the japonica group was divided into two sub-groups, intermediate japonica-cline and japonica /japonica-cline group. Zhe 04A and four japonica×japonica hybrids and their restoring lines fall into the latter group. Meanwhile, the indica group had eight indica type restoring lines of indica×japonica hybrids, three indica control and indica hybrid rice Shanyou 63. When all the twelve combinations were analyzed as a whole, the correlations between the InDel genetic distance and economic yield, number of grains per panicle, grain weight per panicle were significantly positively, but the InDel genetic distance was significantly negatively correlated to seed-setting rate. However, when the eight indica-japonica hybrids were tested only, the InDel genetic distance was only significantly positively correlated with grain weight per panicle and 1000-grain weight. 【Conclusion】Eight combinations from twelve tested Zheyou series combinations were verified attributing to the typical hybrids of japonica sterile line×indica restoring line. Genetic distance tested by InDel molecular marker had higher ability to predict heterosis among different breeding types of hybrids rice, especially for the heterosis of grain weight per panicle. With the increase of InDel genetic distance, the heterosis of tested combinations would be mainly expressed in enhancement of grain weight per panicle.

Genetic Dissection for Kernel Row Number in the Specific Maize Germplasm Four-Rowed Waxy Corn

JIAO Fu-Chao, LI Yong-Xiang, CHEN Lin, LIU Zhi-Zhai, SHI Yun-Su, SONG Yan-Chun, ZHANG Deng-Feng, LI Yu, WANG Tian-Yu

Scientia Agricultura Sinica. 2014, 47(7):  1256-1264.  doi:10.3864/j.issn.0578-1752.2014.07.002

Abstract ( 462 )   HTML ( 2 )   PDF (791KB) ( 934 )   Save

References | Related Articles | Metrics

【Objective】 Kernel row number (KRN) is directly related to maize yield. QTL for KRN was mapped to dissect the genetic basis of KRN, which was helpful to the improvement of yield performance in maize.【Method】In order to dissect the genetic control of KRN of Four-rowed waxy corn, a landrace collected in southern Yunnan, the segregation populations were developed by reciprocally crossing the inbred line derived from Four-rowed waxy corn with Nong531, an inbred with 18-22 rows. The reciprocally crossed F2:3 populations derived from F2 by single seed descend method were separately phenotyped in Beijing (BJ) and Henan (HN). Trials were conducted in randomized complete blocks with two replications. Totally 173 polymorphic SSR primer pairs obtained from the maize genome database (http://www.maizegdb.org) were used to genotype the lines and to construct linkage maps. QTL and their interactions for KRN were detected with ICIM (QTL IciMapping Version3.2 Software) and CIM (QTL Network2.0 Software). The phenotypic variation explained by the major QTL was estimated with SAS GLM software. 【Result】 Phenotypic analysis showed that the average KRN of Four-rowed waxy corn and Nong531 was 4.0 and 19.2, respectively, and the KRN of the reciprocally crossed F2:3 families ranged from 4.0 to 17.4. In two environments, a total of 12 QTL were detected with ICIM, which were located on eight chromosomes. All alleles having negative effect on KRN were from Four-rowed waxy corn. A total of five major QTL could be detected in both environments. qKRN2-1, which was the QTL with the largest effect, explained 18.48% of the phenotypic variance. The other four QTL, which were qKRN4-2, qKRN5-1, qKRN8-2 and qKRN9-1, explained 11.58%, 13.55%, 16.91% and 9.66% of the phenotypic variance, respectively. Five QTL were detected with CIM, four of which were in accordance with the major QTL detected with ICIM, that were qKRN2-1, qKRN4-2, qKRN8-2, and qKRN9-1. Cumulatively, the five major QTL explained 51.5% (BJ) and 54.0% (HN) of the phenotypic variance in the two environments with the GLM model, respectively. Correlation analysis showed that, in the two environments, correlations between the estimated values of phenotypic variance and observed values in F2:3 families were highly significant. In addition, two pairs of QTL located on three chromosomes (Chr. 2, Chr. 4 and Chr. 9) represented epistasis, explaining only a small part of the phenotypic variance, which were 2.90% and 1.80%, respectively. 【Conclusion】 Four-rowed waxy corn possessed a number of alleles with KRN-reducing effect and it could be a good resource in genetic analysis of maize KRN. The five QTL located in bin2.04, bin4.09, bin5.04, bin8.05 and bin9.03 could be considered as the major candidates for fine mapping and cloning in the future.

Evaluation and Analysis of Folic Acid Content in Millet from Different Ecological Regions in Shanxi Province

SHAO Li-Hua, WANG Li, BAI Wen-Wen, LIU Ya-Juan

Scientia Agricultura Sinica. 2014, 47(7):  1265-1272.  doi:10.3864/j.issn.0578-1752.2014.07.003

Abstract ( 608 )   HTML ( 11 )   PDF (591KB) ( 738 )   Save

References | Related Articles | Metrics

【Objective】The contents of folic acid in foxtail millet originated from different regions in Shanxi was analyzed to evaluate the relationship between the variation of folic acid content and ecological distribution in Shanxi, and provide supports for identification of millet quality and breeding.【Method】Totally 245 traditional millet varieties for breeding and planting in Changzhi, Fenyang and Taiyuan were collected, respectively, to determine folic acid content by indirect fluorescent method. The millet seeds were dried at 60℃ after recording the hull colour, and peeled by grinding, and then the color seed grains were marked, and sieved through 100 meshes. Folic acid was extracted by potassium dihydrogen phosphate in waterbath. To obtain the optimum extraction effect, the active carbon adsorbent treated by aniline was added to extraction, the extracts were eluted continuously with a mixture containing 3% ammonia and 70% ethanol, and indirect fluorescent method was used to determine the fluorescence intensity of folic acid oxidized by potassium permanganate.【Result】 The average content of folic acid in millets from Shanxi was 1.53 μg•g-1. Folic acid content in millets exhibited a normal distribution characteristic with the left deviation, indicating that the contents of folic acid in millets were gathered at higher level relative to average. Folic acid content in different regions showed a significant difference. The same variety grown in Fenyang showed the lowest folic acid content compared to that in Taiyuan and Changzhi. Data showed that daily mean temperature, sunshine time and relative humidity had no significant effects on the folic acid content, the rainfall affected significantly the contents of folic acid. The seed colour had no significant impact on folic acid content, but the color of peeled millet grains remarkably affected the contents of folic acid. The millet grains with brown and green color had the highest contents of folic acid, and the grains with yellow, canary yellow, pale yellow and white color were decreased in turn.【Conclusion】It was concluded that folic acid content in foxtail millet germplasms exhibited wide genetic diversity, some foxtail millet varieties with higher folic acid content were identified, thus providing valuable material supports for breeding with high quality and germplasm innovation. The variation range was determined between 0.37 and 2.37 μg•g-1 and the variation coefficient r was 26.2%. Folic acid contents in different ecological regions displayed a significant difference, and the contents of folic acid in millets from late maturing areas were significantly higher than that in mid-maturing areas for spring sowing millets. The color of millet grain significantly affected folic acid content. In this study, total 24 foxtail millet varieties with higher folic acid content were identified, and they accounted for 9.8% of all the tested materials based on the content distribution of folic acid. Among them, Jingu 21, as a major leading cultivar in agricultural production, should be widely extended in Shanxi province, because it is a cultivated variety with higher folic acid content, and the folic acid content in millet variety Jingu 21 reaches 2 μg•g-1.

TILLAGE & CULTIVATION·PHYSIOLOGY & BIOCHEMISTRY·AGRICULTURE INFORMATION TECHNOLOGY

Research Status and Development Discussion on High-Yielding Agronomy of Mechanized Planting Rice in China

ZHANG Hong-Cheng, GONG Jin-Long

Scientia Agricultura Sinica. 2014, 47(7):  1273-1289.  doi:10.3864/j.issn.0578-1752.2014.07.004

Abstract ( 556 )   HTML ( 7 )   PDF (874KB) ( 1349 )   Save

References | Related Articles | Metrics

Rice planting mechanization is a difficult point of the whole-course mechanized development in crop production in China. Based on the reviewing of development situation of rice planting mechanization in China, the high-yielding rules and cultivation techniques of three mechanized planting methods were highlighted and illustrated. (1) Growth characteristics and high-yielding rules of blanket-seedling mechanical transplanting rice grown in floppy disks were generalized systematically. Firstly, population photosynthesis and matter production at the middle and later period were improved by developing appropriate dynamics of stems and tillers and LAI. Secondly, it was the emphasis to increase the effective and efficient biomass accumulation in the middle period, and matter production after heading and the final biological yield. Thirdly, the sufficient population spikelets were composed of enough panicles and larger spike harmoniously, with normal seed-setting rate and 1000-grain weight at the same time. Moreover, the supporting cultivation techniques for high yield were as follows. ① Nursing standardized seedlings for providing biological basis of building starting point of high-yielding population. ② Precise mechanical planting was conducive to the formation of high-yielding population. ③ Promoting tillers growth earlier in the early period could make sure of suitable stems and tillers for high yield slightly before the critical leaf-age for productive tillers. ④ Field draining earlier and lighter, with an appropriate number of peak seedling (1.4-1.5 folds of expected panicle number), could create conditions for reapplying fertilizer of strong stalk and spikelet promotion as early as possible. ⑤ Committed to optimize the growth in the middle period and increase the effective and efficient biomass accumulation, a right amount of population stems and tillers with strong stalk and large panicle was beneficial to constitute a high-efficiency photosynthetic layer. ⑥ Strengthening material production and accumulation for increasing the enrichment of population sink. (2) Many advantages of the high-yielding cultivation of pot-seedling mechanical transplanting rice were also introduced systematically, such as nursing elder seedlings with soil bowl, almost no damage to machine-transplanting precisely, achieving designed basic seedling of high-yielding cultivation accurately, forming an appropriate number of population stems and tillers with strong stalk and large panicle, improving ventilation and light conditions of population, strengthening the resistance to lodging, coordinating source-sink, strengthening strong photosynthetic production in the middle and later periods, making full use of temperature and solar radiation, creating (super) high yield stably and annual yield in the multiple cropping rotation systems, etc. And the key agronomic techniques for high-yielding cultivation of pot-seedling mechanical transplanting rice were as below. ① Nursing age-lengthening seedlings precisely; ② Precise quantitative mechanical planting; ③ Reapplying tillering fertilizer and spikelet- promoting fertilizer appropriately. (3) Characteristics of growth and yield formation of mechanical drilling rice were reviewed, presented with its supporting techniques including determined planting areas reasonably, selecting large-panicle varieties with appropriate growth duration and strong lodging, mechanical sowing earlier and extremely, chemical weed control and fertilizer management. Simultaneously, the existed main problems in the process of rice planting mechanization in China were analyzed further. According to China’s national conditions, the cultivation model raising nurturing standardized seedlings professionally and mechanical transplanting precisely as well as stable high-yielding-high-efficiency agronomic techniques should be the basic direction of mechanized cultivation for the majority of the localities. And mechanical seeding could be applied in several areas with abundant heat of rice season. Then the corresponding research and development (R & D) focuses for mechanized planting of rice were referred. ① High-yielding cultivation of blanket-seedling mechanical transplanting rice should be classified as the main direction of mechanization in the major rice producing areas so as to R & D further. There were several focal points such as enhanced flexibility of blanket-seedling age and seedling quality, straw machine-returning and land preparation and seedling planting precisely, promoting earlier and stably in the field, cultivating a right amount of population stems and tillers with strong stalk and large panicle and increasing population sink. ② Reducing equipment costs, improving operating efficiency and building stable (super) high-yielding-high-efficiency agronomy techniques were the research priorities of pot-seedling mechanical transplanting rice. ③ However, for the mechanical drilling rice, attention should be focused on improving the quality of previous straw machine-returning and land preparation through efficient mechanical operations, and mechanical precise direct seeding and early germinating for achieving expected seedlings. Finally, the technology integration and demonstration of the whole-process mechanized models under the local main mechanized cultivation methods should be done well in accordance with their characteristics in each main region of rice.

Analysis of Salt-Tolerance and Determination of Salt-Tolerant Evaluation Indicators in Cotton Seedlings of Different Genotypes

DAI Hai-Fang, WU Hui, A Man-Gu-Li-?Mai-Mai-Ti-A-Li, WANG Li-Hong, MAI Mai-Ti-?A-Pi-Zi, ZHANG Ju-Song

Scientia Agricultura Sinica. 2014, 47(7):  1290-1300.  doi:10.3864/j.issn.0578-1752.2014.07.005

Abstract ( 619 )   HTML ( 7 )   PDF (710KB) ( 1136 )   Save

References | Related Articles | Metrics

【Objective】As one of the salt-tolerance crops, there is a great difference among different varieties. So, the objective of this study was to investigate the salt tolerance of cotton seedlings of different genotypes which are grown widely in different eras in Xinjiang, to excavate their own salt tolerance genetic resources, to screen suitable evaluation indicators of salt-tolerance, and to establish a mathematical evaluation model for salt tolerance, which would provide a theoretical basis for discovery and promotion of salt tolerant germplasm, breeding of new salt-tolerant varieties and evaluation on salt tolerance of a large number of cotton varieties.【Method】Seventeen varieties that are widely grown at different decades in Xinjiang of China were selected as test materials in this study. Compared to the control, the mass ratio of NaCl to growing medium was 0.006 in salt stress, and the composition of matrix is 1 part grass charcoal plus 1 part vermiculite. After being disinfected and pregermination, cotton seeds of seventeen different genotypes were to sow in the medium. This is helpful to reflect the actual stress environment in field conditions and explain the mechanisms of the tolerance by means of continuous salt stress at seedling stage. The salt comprehensive evaluation of salt tolerant coefficient (STC) about the emergence rate (ER), fresh weight (FW), dry weight (DW) and plant water content (PWC) of each variety, leaf area of the first true leaf (LA), the net photosynthetic rate (Pn), chlorophyll content (Chl) and relative electric conductivity (REC) in leaves of cotton seedlings of different genotypes under salt stress (0.6%) were evaluated by principal components analysis, hierarchical cluster analysis and regression analysis. Physiological performance characteristics of various salt-tolerant types of cotton varieties were analyzed, too.【Result】The results showed that the 11 single indexes in leaves of cotton seedlings of different genotypes under salt stress could be classified into 6 independent comprehensive components by means of principal components analysis. According to the salt-tolerance comprehensive evaluation value (D value) of different cotton varieties, which was calculated from the subordinate function analysis, the 17 cotton cultivars were divided into four salt-tolerant types by cluster analysis, 3 of the 17 varieties were salt-sensitive type, 6 varieties were weak salt-resistance type, 6 were medium salt-resistance type, and 2 were high salt-resistance type. A mathematical evaluation model for cotton salt tolerance was established by means of regression analysis, and D= -1.192+ 0.402REC+ 0.119LA+ 0.274FW+ 0.086Pn+ 1.019Chl (R2= 0.9921). Five indexes closely related to the salt tolerance, LA, REC, FW, Pn and Chl, were screened, too. Evaluation accuracy of all varieties were more than 94.44%, this shows that the five indexes in the regression equation obviously affect the salt resistance of cotton seedlings, and this equation can be used in salt tolerance evaluation of cotton cultivars. Furthermore, the regression analysis and clustering results were mutual authentication. The results showed that, compared with salt-sensitive cotton varieties, REC of cotton leaves of high salt-resistance type was lower than that of other cultivars after treated with 0.6% salt stress, Pn, Chl, LA and FW were higher, LA of high salt-resistant cotton seedlings especially was nearly 2 times larger than other categories varieties.【Conclusion】Higher salt tolerant varieties, which had lighter injury, higher leaf area and photosynthetic capacities under salt stress, were able to maintain higher accumulation ability of salinity and photosynthetic products, decreased ion toxicity from soil, and enhanced salt tolerance of plants. Determination of the five indicators under the same condition can be used for rapid identification and prediction of other cotton varieties, which is very useful for the breeding, promotion, identification and screening of salt tolerant germplasm.

Insect chitin metabolism and plant protection

Insect Chitin Metabolism and Plant Protection

ZHANG Jian-Zhen

Scientia Agricultura Sinica. 2014, 47(7):  1301-1302.  doi:10.3864/j.issn.0578-1752.2014.07.006

Abstract ( 429 )   HTML ( 3 )   PDF (216KB) ( 723 )   Save

References | Related Articles | Metrics

Research Progresses in Insect Glycosyl Hydrolyase Family 20 β-N-acetylhexosamindase

QU Ming-Bo, LIU Tian, CHEN Lei, CHEN Qi, YANG Qing

Scientia Agricultura Sinica. 2014, 47(7):  1303-1312.  doi:10.3864/j.issn.0578-1752.2014.07.007

Abstract ( 468 )   HTML ( 13 )   PDF (674KB) ( 807 )   Save

References | Related Articles | Metrics

β-N-acetylhexosaminidase (Hex), a family 20 glycosyl hydrolyases (GH20), is an essential enzyme for the metabolism of N-acetylhexosamine by catalyzing the removal of β-linked N-acetylhexosamine from the non-reducing ends of glycans, glycoproteins and glycolipids. Insect GH20 β-N-acetylhexosaminidase involves in multiple physiological processes, including chitin degradation, protein N-glycan modification, glycoconjugates degradation and sperm-egg recognition. Because of its important functions, β-N-acetylhexosaminidase may serve as a potential target for designing eco-friendly pesticides. The insect family 20 β-N-acetylhexosaminidase could be grouped into 4 groups according to their phylogenetic relationship and physiological functions, that are Hex1, Hex2, Hex3 and Hex4. Hex1 is mainly expressed in epidermis during insect molting. The RNAi of Hex1 would cause lethal phenotype during insect molting. The old cuticle of the dsHex1 injected larvae could not be shed off. The study of the enzymatic properties of Hex1 demonstrates that it could efficiently hydrolyze β-1,4 linked chitin oligosaccharide with very high specificity but could not act on β-1,3 or β-1,2 linked substrates. It is a special enzyme for chitin degradation. So far, the only crystal structure of insect β-N-acetylhexosaminidase is OfHex1 from Ostrinia furnacalis. The structure at subsite “+1” in OfHex1 explains the high efficiency and specificity of OfHex1 towards chitin oligosaccharides. Hex2 could not be found in insects belong to diptera. It shows highly similarity towards human β-N-acetylhexosaminidase than the other insect β-N-acetylhexosaminidases. The RNAi of Hex2 will cause abnormalities of larval abdomen, pupa and adult appendages. The enzymatic properties of Hex2 show that it is an enzyme with broad substrate-spectrum by hydrolyzing β-N-acetylhexosamine from chitin oligosaccharides, N-glycan and glycolipids. Hex2 may be involved in the degradation of glycoconjugates like the function of human β-N-acetylhexosaminidase. Hex3 is not well studied so far. The RNAi of Hex3 could cause abnormalities during insect molting. It is proved to be in molting fluid and interact with Hex1. The enzymatic activity analysis indicates that OfHex3 is able to degrade chitooligosaccharides, but at a lower rate than that of OfHex1. Besides, Hex3, together with Hex1 and Hex4, are also found to be in the plasma membrane of spermatozoa in dipteran insects, suggesting that it may be involved in sperm-egg recognition. Hex4 is also called FDL because the mutant of this gene will cause fused lobes phenotype of the mushroom body in Drosophila melanogaster. It is another β-N-acetylhexosaminidase with strict substrate specificity. It could exclusively hydrolyze the terminal β-1, 2-GlcNAc residue from the α-1,3 branch instead of the α-1,6 branch of the substrate GnGn. The subcellular localization of Hex4 indicates that it locates in the Golgi apparatus. All these suggest that Hex4 is a β-N-acetylhexosaminidase involved in the modification of N-glycans. So far, although big progress has been achieved on insect β-N-acetylhexosaminidase, only Hex1 has been well studied, including its physiological functions, enzymatic properties and crystal structures. It is proved to be a potential target for designing eco-friendly pesticides. How the other three β-N-acetylhexosaminidases function during insect development and the structure basis of the different enzymatic properties among the four groups of β-N-acetylhexosaminidases are still uncover. This review focuses on the recent progresses on phylogenetic relationship, crystal structure, enzymatic properties and physiological significance of insect β-N-acetylhexosaminidases.

Molecular Characterization and Function of Chitinase 10 Gene (OcCht10) from Oxya chinensis

LI Da-Qi-1, WANG Yan-1, ZHANG Jian-Qin-1, LI Tao-1, SUN Yi-2, ZHANG Jian-Zhen-1

Scientia Agricultura Sinica. 2014, 47(7):  1313-1320.  doi:10.3864/j.issn.0578-1752.2014.07.008

Abstract ( 483 )   HTML ( 2 )   PDF (642KB) ( 9517 )   Save

References | Related Articles | Metrics

【Objective】 The objectives of this study are to obtain cDNA sequence of chitinase 10 gene (OcCht10) from Oxya chinensis, analyze its functional domain and phylogenetic relationship with chitinases from other known insect species, investigate its expression patterns and biological function during molting process, and to provide a new candidate gene for pest control.【Method】 cDNA fragments of OcCht10 were searched from O. chinensis’ transcriptome database. After blast analysis, the cDNA sequence of OcCht10 was assembled and translated, the functional domains of OcCht10 were predicted by bioinformatics methods. Phylogenetic analysis was performed with other insect chitinase 10 amino acid sequences. The first-stranded cDNAs were synthesized by using RNA isolated from integument of each day of 5th instar nymphs and various tissues of the 6th day in 5th instar nymphs. Reverse transcription quantitative PCR (qPCR) was carried out to analyze the gene expression patterns. Biological function of OcCht10 was studied by RNA interference method. The dsRNA primers were designed for dsOcCht10 synthesis in vitro. The dsRNAs were injected into the 2nd day of 5th instar nymphs for RNA interference, integument was dissected for silencing efficiency detection at 24 h after injection by using qPCR method. The phenotype was carefully observed and mortality was calculated till control insects molted to adults.【Result】 The obtained cDNA (9 318 bp) of OcCht10 contained an open reading frame of 8 613 bp, encoding 2 870 amino acid residues and a non-coding region of 705 bp at 3′ end. There were about 500 bp lost in 5′ end. The deduced amino acid sequence included five chitinase catalytic domains and six chitin binding domains. Phylogenetic analysis showed that OcCht10 belonged to chitinase group Ⅱ, the genes from this group were crucial for insect molting based on references. Tissue specific expression analysis of OcCht10 showed that it was predominately expressed in the integument, foregut and hindgut, which developed from ectoderm. The results suggested that OcCht10 may be involved in chitin metabolism of insect integument. Developmental expression patterns showed that OcCht10 was highly expressed before and after molting stages, lower in middle stages of 5th instar nymphs, which implied that OcCht10 could digest chitin of integument during molting process. RNA interference results indicated that the corresponding transcript level was silenced by 70% after OcCht10 dsRNA injection. Compared with the dsGFP injected control group, the nymphs injected with OcCht10 dsRNA displayed slow development and failed to detach old cuticle during molting, the mortality reached 100%.【Conclusion】 The partial cDNA sequence of OcCht10 was obtained from O. chinensis, the mRNA expression of OcCht10 was higher in the integument before molting; OcCht10 is involved in O. chinensis molting process, and dsOcCht10 injection can effectively silence mRNA expression of this gene and result in the block of ecdysis and even death of O. chinensis.

Molecular Characterization and Functional Analysis of Chitin Deacetylase 2 Gene in Oxya chinensis

YU Rong-Rong, DING Guo-Wei, GUO Ya-Ping, MA 恩Bo, ZHANG Jian-Zhen

Scientia Agricultura Sinica. 2014, 47(7):  1321-1329.  doi:10.3864/j.issn.0578-1752.2014.07.009

Abstract ( 463 )   HTML ( 1 )   PDF (627KB) ( 445 )   Save

References | Related Articles | Metrics

【Objective】 In order to provide a theoretical basis for selecting novel target for pest control, molecular characteristics and biological function of chitin deacetylase 2 gene (OcCDA2) from Oxya chinensis were studied.【Method】The cDNA sequences of OcCDA2 were searched from transcriptome of O. chinensis by bioinformatics method, and the conserved domain was anaylzed, the homologous sequences from Tribolium castaneum, Drosophila melanogaster; Anopheles gambiae, Bombyx mori and Choristoneura fumiferana were selected to construct phylogenetic tree. The real-time PCR was applied to detect the relative expression of OcCDA2 in different tissues and developmental stages of the 5th instar nymphs. The RNA interference (RNAi) was performed to study the biological function of the OcCDA2 during molting and development of O. chinensis. 【Result】The full length cDNA sequences of two chitin deacetylase genes were identified. The amino acid analysis showed that they possessed the signal peptide, the open reading frame contained three conserved domains: chitin binding domain (ChBD), low-density lipoprotein receptor chass A domain (LDLa) and chitin deacetylase catalytic domain (CDA). The phylogenetic analysis showed that two variants of OcCDA2 clustered with the CDA2s of five insect species, and the two variants differed only in one exon consisting of about 120 nucleotides, the alternatively spliced regions gathered with CDA2a or CDA2b, respectively. Based on sequence alignment and phylogenetic analysis, two variants of OcCDA2 were named as OcCDA2a and OcCDA2b. Tissue-specific expression analysis indicated that both OcCDA2a and OcCDA2b were mainly expressed in the integument, foregut and hindgut. Developmental expression patterns showed that the relative expressions of OcCDA2a and OcCDA2b were higher in the 1st day of 5th instar nymphs, then decreased gradually, and began to ascend before molting. RNAi results showed that the expression of target genes was significantly reduced in 24 h after dsOcCDA2a or dsOcCDA2b was injected into 1st day of 5th instar nymphs, the silence efficiency of genes was 96.72% and 80.43%, respectively. Compared with dsGFP injected controls, the majority of insects injected with dsOcCDA2 and dsOcCDA2a were unable to shed the old cuticle and led to body warping, finally to death. A minority of nymphs died before molting without obvious phenotypes. A few nymphs could develop to the adults, however, the legs appeared strengthless and the movement became slowly. The mortality rates were 87.5% and 94.4%, respectively. While insects injected with dsOcCDA2b alone could successfully molt to adults, and no phenotype was observed.【Conclusion】The OcCDA2 has two alternatively spliced variants, namely OcCDA2a and OcCDA2b. OcCDA2a plays a key role in the development of O. chinensis. The silence of OcCDA2a leads to the ecdysis failure and death of O. chinensis. However, OcCDA2b is not essential to molting process of O. chinensis.

Expression, Function and Regulation of Chitin Synthase 2 Gene in Locusta migratoria

LIU Xiao-Jian, CUI Miao, LI Da-Qi, ZHANG Huan-Huan, YANG Mei-Ling, ZHANG Jian-Zhen

Scientia Agricultura Sinica. 2014, 47(7):  1330-1340.  doi:10.3864/j.issn.0578-1752.2014.07.010

Abstract ( 550 )   HTML ( 15 )   PDF (903KB) ( 11073 )   Save

References | Related Articles | Metrics

【Objective】Chitin synthase is one of the key enzymes responsible for chitin synthesis in insects. As this enzyme is absent in higher animals, it could be served as a potential target for developing safe and effective insecticides. In our earlier research, the cDNA of chitin synthase 2 gene (LmCHS2, GenBank accession number: GU067731) in Locusta migratoria was cloned. The objectives of this paper are to further study the expression, function and regulation of LmCHS2, and to provide a scientific basis for effective pest control using RNAi methods.【Method】Based on the nucleotide sequence of LmCHS2, a pair of specific expression primers was designed, the expression patterns of LmCHS2 were studied in eggs, nymphs and adults by RT-qPCR. The dsRNA of LmCHS2 was synthesized in vitro, and then injected into the female or male adults on day 1, respectively. The midguts dissected from the injected insects on day 5 were pooled for each RNA extraction. cDNA synthesis and RT-qPCR were performed to determine the down-regulation of LmCHS2. After dissected the whole gut, the midgut changes and integrity of peritrophic matrix (PM) were observed to explore the biological functions of this gene in L. migratoria adults. Locusts were maintained with no food in different times, and feeding again, to observe the changes of guts. Then the transcript levels of LmCHS2 were detected by RT-qPCR. 【Result】 LmCHS2 was almost undetectable during the early and middle embryogenesis, but dramatically up-regulated in late eggs. It was consistently expressed throughout the nymphal and adult stages. After dsCHS2 was injected into the female or male adults on day 1, significantly reduced transcript of LmCHS2 was observed as compared with that of the controls, and resulted in a decreased feeding and a high mortality of insects (78% for female and 85% for male adults). After dissection, it was found that there was virtually no food contained in dsCHS2-injected insects and the average length of midguts and gastric caeca was shorter than that of the control. Furthermore, histological observation of midguts showed that the control locusts contained a fully developed PM, however, locusts injected with dsCHS2 exhibited a disrupted PM or even absence of the PM. Locusts were treated under starvation for 48 h, the midguts hardly contained food and the average length of midguts was significantly shorter than that of the control midguts. From the H & E stained results, it was found that the PM was almost absent in non-fed midguts while the PM of control midguts was well-structured, which was very similar with the RNAi. But after fed again, the insects contained a fully developed PM. When locusts were maintained with no food for 24 h and 48 h, the transcript levels of LmCHS2 were suppressed significantly. When locusts were fed for another 0.5 h period, the transcript levels increased to the control level rapidly, which suggested that feeding affected the expression of LmCHS2. 【Conclusion】LmCHS2 is responsible for chitin biosynthesis of peritrophic matrix of the midgut and plays a key role for the development of L. migratoria. The decreased expression of this gene affected the integrity of the PM, thus hindered the food absorption and led to the mortality of the locusts. In addition, feeding regulated the expression of LmCHS2.

Cloning and Expression of Two Laccase Genes OfLac1 and OfLac2 from the Insect Ostrinia furnacalis

CHEN Peng, QU Ming-Bo, YANG Jun, YANG Qing

Scientia Agricultura Sinica. 2014, 47(7):  1341-1350.  doi:10.3864/j.issn.0578-1752.2014.07.011

Abstract ( 489 )   HTML ( 2 )   PDF (536KB) ( 691 )   Save

References | Related Articles | Metrics

【Objective】 The objectives of this study are to clone two genes encoding OfLac1 and OfLac2 from Ostrinia furnacalis, and to predict physiological functions of these two genes based on the stage-specific and tissue-specific expression patterns of the two genes during development as well as their responses to the ecdysone (20E) treatment. 【Method】 The first fragment of OfLac1 or OfLac2 was amplified by RT-PCR using degenerated primers designed according to the conserved amino acid sequences of the other insects’ Lac1 and Lac2, respectively, and cDNAs generated from RNA isolated from O. furnacalis at pupa stage as template. The 3′ and 5′ regions of OfLac1 or OfLac2 were obtained by RACE using gene specific primers designed according to the first fragment of OfLac1 or OfLac2, respectively. Seven different tissues including epidermis, fat body, trachea, midgut, Malpighian tubule, silk gland and testes were collected from the fifth-instar day-4 larvae of O. furnacalis and the expression levels of OfLac1 and OfLac2 in these tissues were analyzed through semi-quantitative PCR. Twenty-nine samples were collected at different growth periods of O. furnacalis from egg to adult and the expression levels of OfLac1 and OfLac2 were analyzed by using real-time PCR. The fifth-instar day-2 larvae of O. furnacalis were treated with ecdysone (20E) and samples were collected at 1, 4, 8, 12 and 24 h post treatment. The expression levels of OfLac1 and OfLac2 were then analyzed by using real-time PCR. 【Result】 The full-length cDNA of OfLac1 and OfLac2 from O. furnacalis were obtained. The cDNA sequence of OfLac1 gene was 3 065 bp, containing a 5′-untranslated region (5′-UTR) of 222 bp and a 3′-untranslated region (3′-UTR) of 440 bp. It contained an ORF of 2 403 bp encoding 800 amino acid residues with a predicted molecular weight of 90.6 kD and an isoeletric point of 5.34. OfLac2 was 3 405 bp in length, containing a 5′-UTR of 162 bp and a 3′-UTR of 960 bp. The ORF of OfLac2 was 2 283 bp, encoding 760 amino acid residues with a predicted molecular weight of 84.0 kD and an isoelectric point of 6.43. OfLac1 was predicted to contain an N-terminal signal peptide with 22 amino acids in length and OfLac2 was predicted to contain an N-terminal signal peptide with 23 amino acids. The expression pattern analysis revealed that OfLac1 was mainly expressed in midgut and Malpighian tubule, and low amount of OfLac1 transcripts could also be detected in trachea and epidermis. During development, OfLac1 was mainly expressed in adult, and the expression level of OfLac1 during larval and pupal stages was low. OfLac2 was mainly expressed in midgut and silk gland, low amount of OfLac2 transcripts could also be detected in trachea and epidermis. During development, OfLac2 was mainly expressed at prepupa stage during the larval-pupal molting. The expression of OfLac2 was slightly up-regulated in the last day of each instar. The expression levels of OfLac1 and OfLac2 were significantly increased at 24 h after 20E treatment. 【Conclusion】 Two genes encoding OfLac1 and OfLac2 were cloned from O. furnacalis. The expression patterns of OfLac1 and OfLac2 were different, but they were both up-regulated by 20E. OfLac2 was related with the cuticle tanning during molting.

Molecular Cloning, Expression Patterns and RNAi of UDP-N-acetylglucosamine Pyrophosphorylase in Spodoptera exigua

CHEN Jie, CHEN Hong-Xin, YAO Qiong, ZHANG Wen-Qing

Scientia Agricultura Sinica. 2014, 47(7):  1351-1361.  doi:10.3864/j.issn.0578-1752.2014.07.012

Abstract ( 384 )   HTML ( 2 )   PDF (790KB) ( 563 )   Save

References | Related Articles | Metrics

【Objective】 In insects, chitin biosynthesis is a key process for their growth and development. Although it has been studied deeply on fungi, little is known for other genes in the chitin biosynthesis pathway of insects, because most of researches were focused in trehalase and chitin synthase. UDP-N-acetylglucosamine pyrophosphorylase (UAP) is one of the enzymes in the chitin biosynthesis pathway of insects. This paper aims to clarify its effect on the expression of chitin synthase genes and survival rates of Spodoptera exigua. 【Method】 With the technique of molecular biology, such as the polymerase chain reaction (PCR) and RACE technique, the SeUAP from S. exigua was cloned and sequenced. Also, in order to clarify the expression pattern of SeUAP, the total RNA of several tissues of the 2nd day of 5th instars of S. exigua and total RNA of everyday of its life cycle were extracted to detect the expression levels of SeUAP by RT-PCR. Finally, dsRNA was injected into the 1st day of 5th instars of S. exigua to survey the survival rate and detect the expression levels of CHSA and CHSB, which are the last enzymes in the pathway. 【Result】 The complete cDNA sequence of SeUAP was 2 098 bp encoding a protein of 491 amino acid residues, which has a close evolutionary relationship with orthologues in Culex quinquefasciatus, Aedes aegypti, Drosophila melanogaster and Anopheles gambiae. RT-PCR results showed that SeUAP was highly expressed in integument and ovaries. The expression was also detected in trachea and midgut but virtually not in Malpighian tubules and fat body. In addition, RT-PCR results showed that SeUAP was highly expressed at egg stage, L22 (2nd day 2nd instars), L31, L42, L51, P0, P5, P7 and A3, which appeared to be related to the high demanding of chitin synthesis. Further research of RNAi showed that the injection of dsRNA of SeUAP into the larvae with 5 μg caused lethal effect and abnormal phenotypes of pupae, which could be classified into two major groups: (1) Abdomen puparium was formed while head puparium was not and the pupa had difficulty in breaking the old cuticle and getting out; (2) The head puparium was formed and the pupa already broke the old cuticle, but the pupation process stopped and never finished. Silencing of target gene could down-regulate the mRNA levels of chitin synthase gene A (CHSA) and CHSB, especially CHSB. 【Conclusion】 UAP influences gene expression in the chitin biosynthesis pathway and play a role in growth and development of S. exigua.

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

Effects of Different Water and Nitrogen Supplies on Root Growth, Yield and Water and Nitrogen Use Efficiency of Small Pumpkin

LIU Shi-Quan, CAO Hong-Xia, ZHANG Jian-Qing, HU Xiao-Tao

Scientia Agricultura Sinica. 2014, 47(7):  1362-1371.  doi:10.3864/j.issn.0578-1752.2014.07.013

Abstract ( 468 )   HTML ( 2 )   PDF (666KB) ( 849 )   Save

References | Related Articles | Metrics

【Objective】In order to provide a theoretical support for improving the yield and water and nitrogen use efficiency of vegetable crops in view of the questions of irrigation and nitrogen irrationally in the vegetable greenhouse for semiarid northwest area, the responsive mechanism of root growth and distribution to water and nitrogen supply was studied under the different levels of water and nitrogen treatment by the way of exploring the effects of water and nitrogen supply on crop root, yield and water and nitrogen use efficiency.【Method】 The field experiment was conducted with different N-rate and irrigation treatments. Small pumpkin “Jintong” was chosen as a test cultivar. The treatments consist of three different irrigation levels (W3, 1 500 m3•hm-2; W2, 1 100 m3•hm-2; W1, 700 m3•hm-2) and three different nitrogen rate levels (N3, 350 kg•hm-2; N2, 250 kg•hm-2; N1, 150 kg•hm-2). There were 9 treatments in total with randomized block design to study the effects of different water and nitrogen supplies on root growth, yield and water and nitrogen use efficiency of small pumpkin. 【Result】 There are 90% of total root length were concentrated in the 0-40 cm soil layer, the root length density decreased exponentially when the soil layer increased. The root length, yield, water use efficiency (WUE) increased with the improvement of N-rate, and decreased by more nitrogen application at the W1 and W2 irrigation level, but the root length increased with increase of N-rate and there were no significant difference in the yield of small pumpkin at the W3 irrigation level. Compared to N3 treatment, the root length, yield and WUE increased with the increase of irrigation amount, they decreased when the irrigation amount was more than 1 100 m3•hm-2. WUE decreased with the increase of irrigation amount and it reached the highest value (35.59 kg•m-3) in W1N2 treatment. Nitrogen use efficiency (NUE) decreased significantly with the increase of N-rate when the irrigation amount was at W2 and W3 levels, and NUE of N1 and N2 treatment increased significantly compared to N3 treatment. The order of irrigation and nitrogen fertilizer effect on total root length of small pumpkin was nitrogen, water, water and nitrogen interactions. The root length of diameter less than 2 mm presented the parabola trend when N-rate and irrigation increased, and had a better linear regression with the yield of pumpkin. A significant linear relationship between yield and root surface area and the root length of root with the diameter less than 2 mm.【Conclusion】Less or more irrigation and nitrogen fertilizer could reduce the yield, water and nitrogen use efficiency and the characteristic parameters of the roots of small pumpkin. The highest values of yield and characteristic parameters of the roots of small pumpkin were obtained when N-rate and irrigation water amount were 1 100 m3•hm-2 and 250 kg•hm-2, respectively. The water and nitrogen treatments were through the way of root length of root with the diameter less than 2 mm to improve the yield of small pumpkin. It was concluded that the optimal pattern of irrigation and nitrogen fertilization is that the irrigation water amount and N-rate are 1 100 m3•hm-2 and 250 kg•hm-2, respectively.

The Influence of Solar Radiation and Water Demand on Microalgae Production

FENG Ting, Shu GENG, WU Hua-nan, SHI Ping

Scientia Agricultura Sinica. 2014, 47(7):  1372-1378.  doi:10.3864/j.issn.0578-1752.2014.07.014

Abstract ( 376 )   HTML ( 1 )   PDF (556KB) ( 701 )   Save

References | Related Articles | Metrics

【Objective】Microalgae accumulate lipids through photosynthesis, thus can be used to produce biofuel. In addition, microalgae can also be used as supplement for food, oil, and feed. Microalgae cultivation is not compete with terrestrial plants for land resource, thus holding a great potential for the sustainable agricultural development. The commercialization development cannot be obtained without high microalgae productivity. This paper aims to quantify the influence of evapotranspiration and solar radiation on microalgae productivity.【Method】Based on the field production system of Zhaokai Bioenergy Research Center, diatom was cultivated in open pond culture system and the daily productivity data were collected. Meanwhile, the meteorological data were collected from local weather stations of Shenzhen Weather Bureau and the University Town Weather Station. SIMMETEO Model was applied to estimate the daily evapotranspiration data of diatom cultivation system. Based on the photosynthesis and evapotranspiration models, a microalgae productivity model was developed by using solar radiation and evapotranspiration data as inputs.【Result】Results showed that temperature influenced diatom productivity in a relatively small amount with R² about 0.46 in fitted growth curve. Solar radiation as well as evapotranspiration had a significant influence on diatom productivity with R2 above 0.6 in the fitted growth curves. Moreover, the R2 of the sigmoidal regression between daily solar radiation and evapotranspiration was 0.96, which indicates a strong correlation between these two variables. When the daily average solar radiation increased from 9.67 to 12.67 MJ•m-2, evapotranspiration also increased. When daily solar radiation was above 12.67 MJ•m-2, the water demand of algae growth became saturated and maintained at 4.63 mm per day. Evapotranspiration also had a significant influence on microalgae growth with R2 = 0.637 in the fitted growth curves. The cumulative amount of evapotranspiration during the cultivation process represents the total water demand for the growth. Microalgae growth was limited without enough water supplies. Meanwhile, the water demands also increased with the rising productivity.【Conclusion】Optimal solar radiation and evapotranspiration values are essential impact factors for microalgae cultivation. Analyzing the evapotranspiration and water demand during the growth phase will contribute for identifying the important meteorological impacts on algae growth and providing an effective management guide for efficient water application.

HORTICULTURE

Terpenes Biosynthesis Related Gene Transcript Profiles and Terpenes Accumulation of ‘Alexandria’ Grape

SUN Lei-1, ZHU Bao-Qing-2, SUN Xiao-Rong-1, XU Xiao-Qing-3, WANG Xiao-Yue-1, ZHANG Guo-Jun-1, YAN Ai-Ling-1, XU Hai-Ying-1

Scientia Agricultura Sinica. 2014, 47(7):  1379-1386.  doi:10.3864/j.issn.0578-1752.2014.07.015

Abstract ( 482 )   HTML ( 4 )   PDF (749KB) ( 733 )   Save

References | Related Articles | Metrics

【Objective】 The aim of this study was to detect the terpenes and their contents in Alexandria grape, analyze the terpenes accumulation and genes transcript profiles related to their biosynthesis in developing grape berries, to elucidate the regulation of aromatic components for breeding of aromatic grapes. 【Method】 The berries were sampled once a week from veraison to full mature stage, 80-100 berries were randomly chosen, seeds and pedicels were removed, then crushed into juice, after centrifuged, the supernatant was prepared for detection of aromatic substances. Volatile components were extracted by head space solid phase micro extraction method, the compounds were identified by a combination of matching retention index and time with NIST05 library and authentic standards, the compounds were quantified using the internal standard. The primers were designed according to the ORF sequence of DXS gene, RNA of full ripening berries were reverse transcripted into cDNA, which was used as a template to amplify OFR fragments. DXS gene of Alexandria grape was obtained through several times of independent PCR, and single nucleotide polymorphism sites were found by blast of sequence results. Primers of eight key genes in the terpenes biosynthesis pathway were designed, three housekeeping genes of Ubiquitin, EF1-α and GAPDH were used as internal references, transcription abundance of these genes were detected by real-time PCR. 【Result】 The contents of linalool and geraniol were much higher than other monoterpenes, the concentration of various terpenes increased from veraison to ripening, linalool, myrcene and limonene increased by 6-8 times, geraniol, terpineol, geranial and terpinolene increased by 2 to 3 times, rose oxide and neral increased slightly. The concentrations of linalool, gerniol, rose oxide and myrcene were higher than their odor threshold. In the early pathway of monoterpenes biosynthesis, DXS1 gene was gradually up-regulated from veraison and increased by 5 times at the time of 15 weeks after blossom (15WAB). DXS3 gene was up-regulated from 12WAB and DXR gene showed a fluctuating trend, the relative expression level of HDR gene was most abundant in the early pathway and reached 10 to 20 times of DXS1, DXS3 and DXR genes. FPPS gene was up-regulated by 4 times and GPPS by 2 times in the middle pathway, Liner-syn gene and Terp-syn gene were up-regulated from 11 to 15 WAB in the late pathway, and all the genes were down-regulated significantly from 17 WAB. The total contents of monoterpenes accumulated drastically from 12 to 16 WAB, which were in accordance with the expression trend of DXS3, DXR, HDR, GPPS and FPPS genes, the correlation coefficient of total terpenes to DXS3 was 0.831. The ORF length of DXS1 gene was 2 151 bp, it encoded 716 amino acids, 16 single nucleotide polymorphism sites were found by blast of 20 independent sequence results, which caused the change of 4 amino acids.【Conclusion】In the pathway of terpenes biosynthesis, many genes were up-regulated at late-ripening stage, which led to two-eight fold increase of terpenes. DXS3 gene expression was correlated significantly to the total content of terpenes.

Cloning of a Plasma Membrane Aquaporin Gene EjPIP1 in Eriobotrya japonica Leaves and Its Expression Analysis After AM Fungi Inoculation

ZHANG Yan-1, 2 , LI Juan-3, YAO Qing-1, CHEN Jie-Zhong-1, HU You-Li-1

Scientia Agricultura Sinica. 2014, 47(7):  1387-1396.  doi:10.3864/j.issn.0578-1752.2014.07.016

Abstract ( 343 )   HTML ( 1 )   PDF (1005KB) ( 464 )   Save

References | Related Articles | Metrics

【Objective】 The aim of this study was to clone a plasma membrane aquaporin gene EjPIP1 from Eriobotrya japonica variety ‘Zaozhong 6’, and to analyze its sequence characteristics, then to study the effects of AM fungi on the PIP expression pattern and water use efficiency(WUE). 【Method】 A pot experiment with seedlings of loquat cultivar (‘Zaozhong 6’), two treatments (AM and NM) and five replicates were conducted. The leaf water use efficiency was measured by using LI-6400 photosynthesis determination system. The full-length cDNA of EjPIP1 was obtained by the technology of reverse transcription PCR (RT-PCR) and RACE. The cDNA sequence and the deduced amino acid sequence were analyzed by bioinformatics method. The expression patterns of EjPIP1 in leaves and roots by AM fungi inoculation were determined by real-time RT-PCR.【Result】In the effective photosynthetic time(7:00—17:00), the water use efficiency of loquat leaves were significantly increased by AM fungi inoculation. Loquat plasma membrane aquaporin gene EjPIP1 (GenBank Accessin No. JX041627) was cloned. The full-length cDNA of EjPIP1 gene consists of 1 142 bp and contains a 861 bp open reading frame (ORF) encoding 286 amino acid proteins. Two highly conserved NPA (Asn-Pro-Ala, asparagine-proline-alanine) motifs of aquaporin were identified in EjPIP1. The homologue analysis revealed that the amino acids sequence of loquat PIP1 was highly homologous with other species, especially apple (Malus domestica), peach (Prunus persica) and strawberry (Fragaria chiloensis), which was up to 97%, 92% and 90%, respectively. Real-time PCR analysis revealed that EjPIP1 could be expressed in different loquat tissues including root and leaf, and the relative expression in leaf was higher than that in root. Furthermore, under well-watered condition, the expression of EjPIP1 in loquat leaf was significantly down-regulated by AM fungi inoculation, while the expression of EjPIP1 in root was up-regulated. 【Conclusion】 The water use efficiency of loquat leaves was significantly increased by AM fungi inoculation. A plasma membrane intrinsic proteins (PIPs) gene, designated as EjPIP1, was cloned from leaf of loquat (E. japonica ‘Zaozhong 6’). And the expression of this corresponding gene in loquat leaves and roots were influenced by AM fungi, which helped to improve water use efficiency of loquat seedlings after being inoculated with AM fungi.

STORAGE·FRESH-KEEPING·PROCESSING

Effect of Different Drying Methods on Drying Efficiency and Quality of Purple Sweet Potato

LI Wen-Feng, XIAO Xu-Lin

Scientia Agricultura Sinica. 2014, 47(7):  1397-1408.  doi:10.3864/j.issn.0578-1752.2014.07.017

Abstract ( 554 )   HTML ( 2 )   PDF (630KB) ( 1333 )   Save

References | Related Articles | Metrics

【Objective】In order to improve the drying efficiency and drying quality of purple sweet potato (PSP), the effect of different drying methods on moisture lost, value of color difference, anthocyanins, polyphenol and antioxidant of PSP were studied.【Method】The PSP were dried by hot air cross-flow drying, forced air drying, air-impingement jet drying and hypoxia air-impingement jet drying. Firstly, the effect of the three kinds of drying methods including hot air cross-flow drying, forced air drying and air-impingement jet drying on drying curves, drying rate curves and effective moisture diffusion coefficient under the conditions of drying temperature of 70℃, slice thickness of 1.93 mm and microwave pre-treatment for 3 min was studied. Secondly, the effect of four kinds of drying methods including hot air cross-flow drying, forced air drying, air-impingement jet drying and hypoxia air-impingement jet drying on color, total anthocyanins content, total polyphenol content and DPPH• scavenging activity of PSP was analyzed under the conditions of drying temperature of 70℃, slice thickness of 1.93 mm and microwave pre-treatment for 3 min. Furthermore, the influence of four factors including air temperature, air velocity, nozzle distance and slice thickness of hypoxia air-impingement jet drying on the color, total anthocyanins content, total polyphenol content and DPPH• scavenging activity of PSP were studied. 【Result】According to the experimental data, there was a similarity law in water loss of PSP, compared with the most results of food material drying test. Forced air drying, hot air cross-flow drying and air-impingement jet drying of PSP mainly occurred in the falling rate drying period. There was no stage of constant rate drying in the drying process. The drying rate of air-impingement jet drying was 84.04% higher than the forced air drying, and it was also 61.60% higher than hot air cross-flow drying. The moisture content of PSP decreased very fast during the first 40 min of air-impingement drying process. But the moisture content of PSP decreased very slowly in the later drying process. Effective moisture diffusion coefficient of forced air drying, hot air cross-flow drying, air-impingement jet drying was 9.62×10-9, 10.23×10-9, and 15.02×10-9 m2•s-1, respectively. In this research, the total anthocyanins content, total phenols content and DPPH• scavenging activity of the PSP sample was 90.85 mg•100g-1 and 262.14 mg•100g-1, respectively. Compared to the forced air drying, hot air cross-flow drying and air-impingement jet drying, the color was better and the total anthocyanins content, total phenols content and DPPH• scavenging activity were higher by hypoxia air-impingement jet drying. The total color difference, total anthocyanins content, total phenols content and DPPH• scavenging activity of hypoxia air-impingement jet dried PSP was 20.35, 34.79 mg•100g-1, 139.26 mg•100g-1 and 28.49%, respectively. In the experiments of the effect of different hypoxia air-impingement jet drying conditions on quality of dried PSP, with the drying temperature increase and air velocity, nozzle distance and slice thickness decrease, the total anthocyanins content, total phenols content and DPPH• scavenging activity decreased, but the value of color difference reduced. In addition, the highest survival rate of total anthocyanins content, total phenols content and DPPH• scavenging activity of dried PSP was 59.58%, 82.35% and 82.05%, respectively.【Conclusion】The drying efficiency and quality of air-impingement jet drying PSP is higher than the forced air drying and hot air cross-flow drying. In addition, the hypoxia air-impingement jet drying can improve the dried sample quality of PSP based on the normal air-impingement jet drying.

ANIMAL SCIENCE·VETERINARY SCIENCERE·SOURCE INSECT

Study on Marginal Profits of Traits in the Breeding Goal in Sanhe Cattle

MA Qiu-Meng-1, 2 , QIN Chun-Hua-1, 3 , WU Hong-Jun-4, LIU Ai-Rong-5, WANG Ya-Chun-1, SHI Yuan-Gang-2, ZHANG Sheng-Li-1

Scientia Agricultura Sinica. 2014, 47(7):  1409-1416.  doi:10.3864/j.issn.0578-1752.2014.07.018

Abstract ( 378 )   HTML ( 3 )   PDF (523KB) ( 611 )   Save

References | Related Articles | Metrics

The Sanhe cattle is a domestic dual purpose breed formed and approved in China in 1983, which was named after its origin of Sanhe region in Hulunbuir pasture land. Sanhe cattle is famous for its durability, suitability for grazing, cold resistance and strong adaptability. This study was conducted to estimate the marginal profits of traits in breeding goal and provide a base for designing breeding plan in Sanhe cattle. 【Method】 The performance, economic and nutrition parameters of Sanhe cattle were obtained according to theoretical nutrition demand of dairy and beef cattle, as well as the production and breeding data in 2011 from Xiertala Cattle Breeding Farm in Inner Mongolia, China, which are the central production region of this breed. Also, the breeding objective traits were proposed based on the production and breeding system of Sanhe cattle. A bio-economic profit model was fitted using Profit Equation method, and the marginal profits were calculated for the milk performance, beef performance and functional traits, then the sensitivity analysis was run. 【Result】The results indicated that ten breeding objective traits for Sanhe cattle included three milk performance traits (milk yield, fat percentage, and protein percentage), three beef performance traits (birth weight, fattening daily gain, and dressing percentage), and four functional traits (somatic cell count, age at first calving, calving interval, and productive life). The total profit of Sanhe cattle was RMB 4 808.26 with a fixed scale of the population. The profit generated by milk production, culling cows and fattening bulls account for 67.08%,7.8% and 20.58% of the total profit, respectively. For the cost, the variable cost accounts for 96.89% of the total cost. Feeding cost is the main factor affecting the variable cost. The feeding cost of the heifers, milking cows and fattening bulls account for 45.69%, 20.46%, and 13.15% of the total cost, respectively. When raw milk price considering milk compositions and quality (fat percentage, protein percentage, and somatic cell count), the marginal profits of objective traits in Sanhe cattle were RMB 2.26 (milk yield, MY, kg), RMB 40.48 (fat percentage, FP,%), RMB 61.39 (protein percentage, PP,%), RMB -231.52 (somatic cell count, SCC), RMB 6.19 (birth weight, BW, kg), RMB 1.39 (fattening daily gain, FDG, g/day), RMB 76.70 (dressing percentage, DP, %), RMB 0.25 (productive lifetime, PL, day), RMB -1.62 (age at first calving, AFC, day), and RMB -2.71 (calving interval, CI, day)，respectively. The relative economic weights for milk performance, beef performance and function traits is 62﹕14﹕22, closing to 3﹕1﹕1. With the increase of 10% of the milk price, the marginal profits of MY and CI increased by RMB 0.29, RMB 0.17, respectively; with the increase of 10% of the beef price, the marginal profits of BW, FDG, DP and CI increased by RMB 0.81, RMB 0.15, and RMB 7.86, respectively and PL decreased by RMB 0.03; with the increase of 10% of the feed price, the marginal profits of MY, FP, PP, BW, AFC, FDG decreased by RMB 0.05, RMB 0.58，RMB 0.34, RMB 0.21, RMB 0.16, and RMB 0.01, respectively and CI and PL increased by RMB 0.88 and RMB 0.06, respectively. 【Conclusion】Change of market price would largely affect the marginal profits of milk and beef production in Sanhe cattle. The bio-economic profit model derived from this study will provide a reference for dual purpose cattle breeding in China and a basis for development of breeding goals for Sanhe cattle in the future.

The Preparation of mRNA Pluripotency Transcription Factors and Its Expression in Xuhuai Goat Fibroblasts

QIU Feng-Long, ZHANG Ya-Ni, NI Rong, LI Wei, CHEN Ting-Feng, WEI Guang-Hui, LI Bi-Chun

Scientia Agricultura Sinica. 2014, 47(7):  1417-1426.  doi:10.3864/j.issn.0578-1752.2014.07.019

Abstract ( 500 )   HTML ( 1 )   PDF (987KB) ( 519 )   Save

References | Related Articles | Metrics

【Objective】 The genes of Oct4, Sox2, Klf4 and c-Myc were cloned, the pMD19-T-oct4, pMD19-T-sox2, pMD19-T-klf4, and pMD19-T-c-myc recombinant plasmids were constructed, and then the pcDNA3-Oct4, pcDNA3 -Sox2, pcDNA3-Klf4 and pcDNA3-c-Myc recombinant plasmids containing T7 promoter were constructed. According to the in vitro transcription, the four mRNA transcription factors were obtained, and it was made to express stably in Xuhuai goat fibroblasts. 【Method】Through the testicular, skin and small intestine tissue, the Oct4, Sox2, Klf4, and c-Myc genes coding sequence were obtained by RT-PCR, and were cloned into pMD19-T vector, and then cloned into the eukaryotic expression pCDNA3.0 vector that contains T7 promoter, and then the pcDNA3-Oct4, pcDNA3-Sox2, pcDNA3-Klf4 and pcDNA3-c-Myc recombinant plasmids were constructed. Recombinant plasmids were linearized using XhoI and XbaI, in vitro transcription of the transcription factors was made to obtain the mRNA using the instructions of in vitro transcription, the mRNA was detected, its stability and concentration were determined, Lipofectamine 2000 transfection was carried out according to the proportion of Lipofectamine﹕mRNA=1﹕1. Western blot technique and immunofluorescence assay were used to detect the mRNA expression in Xuhuai goat fibroblasts 24 h post-transfection. 【Result】 The length of Xuhuai goat Oct4, Sox2, Klf4 and c-Myc genes coding sequence were 1 083 bp, 962 bp, 1 434 bp and 1 320 bp, and confirmed by TA cloning, the sequences similarity of coding sequences of sheep, human, cattle and pig were above 89%. The mRNA of four transcription factors by transfection to Xuhuai goat fibroblasts were localized in the nucleus. The mRNA of four transcription factors expression in fibroblasts proteins was consisted with the expected size, 38 kd, 34 kd, 50 kd and 48 kd. 【Conclusion】The Xuhuai goat Oct4, Sox2, Klf4 and c-Myc transcription factors were cloned successfully, and the four mRNA can express in fibroblasts stably, thus a foundation for further study of the function of the genes Oct4, Sox2, Klf4, c-Myc and goat somatic cell reprogramming is established.

Evaluation of the Biological Safety of Phytosterols for Broilers

WANG Li-Wen-2, JIANG Yong-1, JIE Jing-Jing-1, 吕Lin-1 , ZHANG Li-Yang-1, SUO Hai-Qing-1, LUO Xu-Gang-1

Scientia Agricultura Sinica. 2014, 47(7):  1427-1437.  doi:10.3864/j.issn.0578-1752.2014.07.020

Abstract ( 361 )   HTML ( 2 )   PDF (1280KB) ( 773 )   Save

References | Related Articles | Metrics

【Objective】An experiment was conducted to investigate the effect of dietary phytosterol level on growth performance, hematology, plasma biochemical traits, weight or length indices of organs, and histopathological changes of Arbor Acres broilers during periods of days 1-21 and 22-42 to evaluate the biological safety of phytosterols for broilers chicks. 【Method】A total of 240 1-d-old broilers (half male and half female) were randomly allocated by bodyweight to one of four treatments with six replicate cages of ten broilers (half male and half female) per cage. Birds were fed the phytosterol-unsupplemented diet with background about 300 mg•kg-1 of total phytosterol or the basal diet supplemented with 80, 400, or 800 mg•kg-1 phytosterols for an experimental duration of 42 days, and which was divided into two periods: start period from day 1 to 21 and growth period from day 22 to 42. Data from the experiment were analyzed by the general liner model procedure of SAS 9.0. 【Result】 Results showed that body weight gain, feed intake, feed conversion and mortality from the starter period and weight gain, feed conversion and mortality from growth period were not affected (P＞0.05) by the addition of phytosterols in diets, but birds fed the diet supplemented with 400 mg•kg-1 phytosterols had lower (P＜0.05) feed intake during growth period compared with the control group or the group supplemented with 80 mg•kg-1 phytosterols; most of biochemistry parameters in chicks from start period and all of biochemistry parameters from growth period were not affected by the addition of phytosterols in diets in the growth period, but which affected the concentration of total protein, album, and creatine, and alanine transaminase activity in plasma on day 21, and the total protein and album concentrations and alanine transaminase activity in plasma on day 21 increased (P＜0.05) when birds were fed the diet supplemented with 800 mg•kg-1 phytosterol. The addition of 400 mg•kg-1 phytosterol reduced (P＜0.05) the plasma creatine concentration on day 21. The relative weight of organ was not affected (P＞0.05) by the addition of phytosterols in diets except for pancreas, and the relative weight of pancreas on day 42 was significantly (P＜0.05) increased by the addition of 80 or 400 mg•kg-1 phytosterols. There were no significant differences (P＞0.05) in blood physiological among all treatment groups in the experimental period. No pathological changes in heart, liver, lung, kidney, and pancreas were observed among all phytosterol treatment groups at the end of the experiment. 【Conclusion】 Phytosterols added in diets at 80 to 800 mg•kg-1 have no significant effect on growth performance, hematology, plasma biochemical traits, and the development of organs, and there are no toxic response in broilers. So when corn-soybean meal basal diet containing 300 mg•kg-1 of phytosterols, the upper limitation of added phytosterols at 80 mg•kg-1 (containing about 400 mg•kg-1 total phytosterols ) in the corn-soybean meal diet would be safe for broilers.

Pharmacokinetic Interaction of Ivermectin and Triclabendazole Following Co-administration to Sheep

GAO Yan-Yan-1, BA Ya-尔2, HA Si-Su-Rong-1

Scientia Agricultura Sinica. 2014, 47(7):  1438-1444.  doi:10.3864/j.issn.0578-1752.2014.07.021

Abstract ( 543 )   HTML ( 1 )   PDF (524KB) ( 600 )   Save

References | Related Articles | Metrics

【Objective】 The objective of this study is to illustrate the pharmacokinetic characteristics and interactions of ivermectin and triclabendazole in helminth infected sheep following co-administration. 【Method】 A total of 15 Ordos merino sheep in lactation period of 4 years old, average body weight of (39.3±3.2) kg, naturally infected with gastrointestinal helminth (EPG≥1500) were selected by using McMaster’s method. The testing sheep were randomly divided into 3 treatment groups of 5 sheep in each group. The sheep in groupⅠ were subcutaneously injected with IVM alone (0.2 mg•kg-1), that in groupⅡ were orally administered with TCBZ alone (15 mg•kg-1), and that in groupⅢ were combined administrated with TCBZ (15 mg•kg-1 po.) and IVM (0.2 mg•kg-1, sc.). 5 mL of blood samples were collected from the jugular vein from 0.75, 2, 4, 8, 16, 24, 36, 48, 72, 120, 192 and 336 h after administration, and the concentrations of IVM, TCBZ and TCBZSO in each sample were detected by high-performance liquid chromatography (HPLC) with fluorescence detection and UV detection, respectively. Chromatographic conditions: a reversed-phase C18 column, InertsilODS-SP (5 µm, 4.6×150 mm, I.D.); mobile phase of V (methanol and acetonitrile)﹕V (water) =95﹕5; excitation wavelength of 364 nm, emission wavelength is 470 nm; the flow rate was 1.0 mL•min-1; column temperature was at room temperature; the injection volume was 20 µL. The pharmacokinetic characteristics were calculated by Phoenix WinNonlin using non-compartmental analysis (NCA) model. 【Result】The results showed that the chromatographic peaks of IVM and internal standard AVM were well separated with retention time of 8.73 min and 6.16 min, which were not influenced by the other interfering peaks of plasma. The retention time of TCBZ, TCBZSO and internal standard of mebendazole were 10.04 min, 5.53 min and 3.42 min, respectively, and well seperated with no interfering peaks. Therefore, the methods established in this study are feasible to detect different targets in testing samples. For IVM, peak plasma concentration (Cmax) and area under the concentration time curve (AUC) were decreased significantly, however, the apparent volume of distribution (Vd), elimination half-life (T1/2ke) and body clearance (CLb) were increased significantly following combined administration of IVM and TCBZ. For TCBZSO, Vd, mean residence time (MRT) and Tmax were significantly increased, however, T1/2ke was significantly decreased in co-administration group. 【Conclusion】In conclusion, the ivermectin and triclabendazole was pharmacokinetically interacted each other following combined use in helminth infected sheep and sufficient attention should be paid to co-administration of them in clinical practices. In order to ensure the full play of the original drug efficacy and synergistic action, the blind compatibility and combination of drugs only based on the in vitro pharmacodynamic synergism should be avoided.