Scientia Agricultura Sinica

Fine Mapping and Physiological Characteristics of A Green-Revertible Albino Gene gra75 in Rice

WANG Ping-Rong, WANG Bing, SUN Xiao-Qiu, SUN Chang-Hui, WAN Chun-Mei, MA Xiao-Zhi, DENG Xiao-Jian

Scientia Agricultura Sinica. 2013, 46(2): 225-232. doi:10.3864/j.issn.0578-1752.2013.02.001

Abstract ( 745 )

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【Objective】 On the basis of phenotypic and physiological characteristics of a green-revertible albino mutant gra75, the mutant gene was finely mapped. 【Method】A green-revertible albino mutant was isolated from ethyl methanesulfonate mutagenesis. Phenotypes of the mutant were observed, and its physiological characteristics and main agronomic traits were analyzed. Genetic mapping of the mutant gene was conducted by using molecular marker and F2 mapping population of gra75/Zhefu 802 to identify this candidate gene. 【Result】 Under paddy ?eld conditions, gra75 mutant plants exhibited a normal green phenotype up to the third-leaf stage. Subsequently, it produce albino phenotype during the fourth to seventh leaf stage, but then it developed nearly normal green leaves from the eighth-leaf stage. At the mature stage, the gra75 mutant and its wild-type had no significant difference in major agronomic traits. On the other hand, chlorophyll and carotenoid content in albino leaves of the mutant obviously decreased at seedling stage. Electron microscopic observation revealed that the number of chloroplasts, thylakoid membranes, granums and starch granules significantly decreased in mesophyll cells of the albino leaves. Genetic analysis suggested that the mutant phenotype of gra75 was controlled by one recessive nuclear gene. The gra75 mutant gene was mapped between InDel markers HC1 and HC2 on the short arm of rice chromosome 6, with genetic distances of 0.06 cM and 0.6 cM, respectively, and with a physical interval of 120 kb. By analyzing and sequencing the candidate genes in this genomic region, it was found that there was a single nucleotide change (C716T) in exon 5 of LOC_Os06g07210 gene encoding RNRL1 (ribonucleotide reductase large subunit) in the gra75 mutant, which caused a missense mutation (Ala to Val) in the encoded product.【Conclusion】The gra75 mutant gene was allelic to V3 (Virescent3) gene. However, the green-revertible albino character of gra75 mutant could express steadily at seedling stage, and moreover, this albino phenotype did not affect significantly main agronomic traits at the mature stage. So the mutant gene of gra75 could be applicable in hybrid rice breeding as a leaf color marker gene.

Genetic Diversity and Association Analysis Using SSR Markers in Barley

LAI Yong, WANG Peng-Xi, FAN Gui-Qiang, SI Er-Jing, WANG Jin, YANG Ke, MENG Ya-Xiong, LI Bao-Chun, MA Xiao-Le, SHANG Xun-Wu, WANG Hua-Jun

Scientia Agricultura Sinica. 2013, 46(2): 233-242. doi:10.3864/j.issn.0578-1752.2013.02.002

Abstract ( 864 )

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【Objective】In order to provide useful information for hybridization combination of parent materials and molecular assisted breeding, the genetic diversity of parent materials was determined by using SSR markers and their association with some agronomic traits were detected. 【Method】 A total of 86 SSR markers were screened for polymorphism among parent materials, and then the analysis on genetic diversity of these materials were detected. Population structure was analyzed through 57 selected markers, and then association analysis between SSR markers and 5 agronomic traits were performed using TASSEL GLM (General Linear Model) and MLM (Mixed Linear Model) programs.【Result】A total of 200 alleles were found from 113 materials, ranged from 1 to 5. Allele frequency ranged from 0.0088 to 1.0000 and Shannon index ranged from 0.0000 to 1.2236. The genetic similarity ranged from 0.5504 to 0.9897, with the mean of 0.7477. Genetic structure analysis showed that the population of these parent materials was composed of 4 subpopulations. There were 9 SSR markers associated with plant height, spike length, awn length, grain number per spike and Spikelet Compactness under GLM program, and the rate of explanation on the phenotype of related marker ranged from 0.0507 to 0.2766. There were 6 SSR markers associated with plant height, awn length and spikelet compactness under MLM program, and the rate of explanation on the phenotype of related marker ranged from 0.0238 to 0.1999.【Conclusion】Genetic diversity and population structure of 113 materials were analyzed through SSR markers and their association with phenotypes were detected. Nine markers, associated with plant height, spike length, awn length, grain number per spike and spikelet compactness, were determined under GLM program. And 6 markers associated with plant height, awn length and spikelet compactness were determined under MLM program. These markers were on chromosomes 1H, 2H, 3H, 4H and 7H, respectively.

Effects of 2A Type Protein Phosphatase on the Development of Setosphaeria turcica

SHEN Shen, WANG Jing-Jing, HAO Zhi-Min, LI Po, LI Zhi-Yong, SUN Zhi-Ying, HAO Jie, TONG Ya-Meng, DONG Jin-Gao

Scientia Agricultura Sinica. 2013, 46(2): 243-249. doi:10.3864/j.issn.0578-1752.2013.02.003

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【Objective】 The aim of this research is to preliminarily explore the function of type 2A phosphoprotein phosphatase (PP2A) in regulating the development and pathogenicity of S. turcica with the usage of cantharidin, the specific inhibitor of PP2A, and lay a theoretical foundation in researching the new antifungal agents and the prevention of plant diseases. 【Method】 S. turcica was treated with cantharidin at different concentrations to observe and investigate colony growth, conidial yield, conidial germination, appressorium production, melanin biosynthesis and HT-toxin activity of the pathogen. 【Result】 The higher the concentration of cantharidin was, the more strongly the hyphal growth of S. turcica was inhibited. The diameter of 8-day-old colony on the PDA media with cantharidin of 160 μmol•L-1 was 41.7% of that of the mock. Meanwhile, the conidial production in treated groups was higher than that of the CK. When the concentration of cantharidin was up to 160 μmol•L-1, the conidial production was 15.5 times as many as that of the CK. The results of the influences of cantharidin on conidial germination, appressorial formation and invasion showed that the sensitivity of these developmental stages from high to low was appressorial formation > conidial germination > appressorial invasion. Furthermore, the contents of intracellular melanin in treated groups were all higher than that of the CK which was 0.13 g•L-1, and the content of intracellular melanin was positive related with the concentration of cantharidin. 【Conclusion】Cantharidin could significantly inhibit colony growth, conidial germination, appressorium production of S. turcica and promote conidiation and the biosynthesis of intracellular melanin.

Discussion of Naming for Two Subspecies of Oryza sativa L.

LING Qi-Hong, ZHANG Hong-Cheng, DING Yan-Feng

Scientia Agricultura Sinica. 2013, 46(2): 250-256. doi:10.3864/j.issn.0578-1752.2013.02.004

Abstract ( 765 )

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The naming method of O. sativa L. subsp. indica Kato and O. sativa L. subsp. japonica Kato for O. sativa L. proposed by Kato was used internationally at the present, which was inexplicable in fact. In this paper, a multitude of naming methods of O. sativa L. subsp. were analyzed and compared. The classification of O. sativa L. subsp. hsien Ting and O. sativa L. subsp. keng Ting was scientific and appropriate, being worthy of uniform application.

Relationship Between the Source and Sink of Spring Maize with High Yield

WANG Yong-Hong, WANG Ke-Ru, ZHAO Ru-Lang, WANG Kai, ZHAO Jian, WANG Xi-Mei, LI Jian, LIANG Ming-Xi, LI Shao-Kun

Scientia Agricultura Sinica. 2013, 46(2): 257-269. doi:10.3864/j.issn.0578-1752.2013.02.005

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【Objective】 Clearifing the relationship between the source and sink characteristics and the quantitative traits of spring maize with high yield (15 000 kg•hm-2) and to reveal the regulation of the source, sink and yield at different densities.【Method】Four experimental sites in maize regions with high yield of Xinjiang and Ningxia of China with similar climatic characteristics were set up (The 71st groups of the Fourth Agricultural Divisions of Xinjiang, the Qitai Farm of the Sixth Agricultural Division of Xinjiang, the experimental farm of Ningxia University, the Tongxin County of Ningxia). The resistant, density tolerance, high-yielding variety Zhengdan-958 was planted in accordance with the uniform cultivation and management practices. The treatments include 12 densities (15 000 to 180 000 plant/hm2) in order to create different types of source and sink, different yield levels (＞15 000 kg•hm-2), the characteristics of the material production of maize, the maximum leaf area index (LAI), photosynthetic potential, grain weight per plant, single-grain number, harvest index, the ratio of grain to leaf, sink features, and their interrelationship.【Result】The high-yield maize dry matter accumulation and grain yield showed a quadratic function, 15 000 kg•hm-2 maize dry matter accumulation was 24 937-54 895 kg•hm-2, 19 270 kg•hm-2 (the highest yield) of dry matter accumulation of 37 417 kg•hm-2, pre-anthesis and post-anthesis accounted for 44.31% and 55.69%, respectively. The high yield maize production and maximum LAI, LAD showed a quadratic function relationships. The maximum LAI of the above 15 000 kg•hm-2 was 3.9-11.4, photosynthetic potential was 113 401-502 703 m2•d, LAI and LAD of the highest yield of maize were 6.88 and 348 142 m2•d. The yield and ear weight per plant had a negatively significant correlation (r =0.7188**), ear weight per plant of maize with the yield above 15 000 kg•hm-2 was 95.5-289.6 g, the highest yield of ear weight per plant was 169 g, yield and kernel number showed a function of quadratic relationship of maize with 1 5000 kg•hm-2 yield, the kernel number was 366.6-545.9. The kernel number of the highest yield of maize was 469. The yield and weight of kernel showed a quadratic function relationships. The 1000-kernels-weight was 232.6-388.6 g, the 1000-kernel-weight of maize with the highest yield was 361.0 g. The high yield maize production and harvest index had a quadratic function relationship, the harvest index was 31.5-61.9%, while that of the highest yield was 51.5%. The yield and kernel number/leaf area and kernel number/leaf dry weight was not significantly correlated (rkernel number/leaf area=0.1520, rkernelr/leaf dry weight =0.2577), the kernel weight/leaf dry weight was significantly (r=0.5847*) and had a quadratic relationship, the kernel weight/leaf weight was 1.1-7.13, kernel weight/leaf area was 149.4-506.5 g•m-2, the highest yield of kernel-leaf ratio was 5.39 and 366.4 g•m-2 of maize with yield above 15 000 kg?hm-2.【Conclusion】At different yield levels, the proportion of dry material accumulation and the photosynthetic potential existed difference in pre-anthesis and post-anthesis, the higher yield needs higher dry matter accumulation and photosynthetic potential, and the higher dry matter accumulation at pre-anthesis, the higher the proportion of photosynthetic potential at post-anthesis, As Zhengdan-958 is a type of source deficiency, which is a main limiting factor of production in low-density, so increasing the density could improve yield. The primary mechanism of yield is increase of leaf source. The high-density conditions, the source and sink increased at the same time but the increased proportion of the relative lack of sink was the main limiting factor of production, the main mechanism of yield increase improved seed setting rate and increased grain weight by the sink expansion methods.

Behavioral Differences of Ethanol Extracts from Sprouting Shoot Tips of Different Sweetpotato Varieties in the Process of DPPH• Scavenging Reaction

XIE Xiao-Huan, ZHAO Ying, LUO Wei, XU Sen, ZENG Ling-Jiang, YANG Chun-Xian, FU Yu-Fan

Scientia Agricultura Sinica. 2013, 46(2): 270-281. doi:10.3864/j.issn.0578-1752.2013.02.006

Abstract ( 559 )

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【Objective】Sweetpotato[Ipomoea batatas(L.)Lam.] vine is composed of 3 parts, namely leaf, petiole and stem. It has received more and more attentions from the researchers, consumers, and is popular in vegetable market because sweetpotato vine is an excellent source of antioxidant and is superior in its physiological health care functions to other commercial vegetables. The indexes reflecting the reaction of 1,1-diphenyl-2-picrylhydrazyl (DPPH•) scavenging of ethanol extracts from sprouting shoot tips of 65 sweetpotato clones (including 16 varieties)were investigated to explore these sweetpotato clones behavioral differences in the DPPH• scavenging reaction process.【Method】The DPPH• scavenging rates at 19 moments between 0 and 210min of the reaction duration among the ethanol extract concentration of 0.1, 0.2, 0.4, 0.8, 1.6, 2.0 mg•mL-1 from sprouting shoot tips of 65 sweetpotato clones were determined, and the reaction rate and its change, EC50 and the corresponding times spent for scavenging rate reaching 50%, 80% in all concentrations were calculated, and 65 clones were classified by cluster analysis according to these indexes.【Result】①Although behaviors of ethanol extract in their DPPH• scavenging process performed continuity among 65 clones, these clones could still be classified into five types, and types Ⅱ, Ⅲ accounted for the majority. ② The reaction rates in 0—1 min and 1—3 min, scavenging rates at 210 min under all concentrations of type Ⅰ including one clone significantly less than those of other types. And its scavenging rate could not achieve 50% even the concentration of ethanol extract increased to 2.0 mg•mL-1. ③The average antioxidant capacity of type Ⅱ with 44 clones was significantly better than that of type Ⅰ and inferior to those of other types. Only the concentration of ethanol extract reached to 0.8 mg•mL-1, scavenging rates of 24 clones in typeⅡ could attain to 50%, but that would spend 83.89 min, and none could achieve 80%. When the concentration increased to 2.0 mg•mL-1, 37 clones in typeⅡ could achieve 80% after 59.23 min. ④The scavenging rate variation curves of type Ⅲ consisting of 17 clones, scavenging rates at 210 min, EC50, reaction rates, the proportion of the number of clones which scavenging rates could reach 50% and 80% and corresponding times spent, were all between type Ⅱ and Ⅳ and deviated to the latter. ⑤Type Ⅳ contained one clone and type Ⅴ included 2 clones. EC50 of both types were lower than those of the former three types, and their scavenging rates could achieve 50% even at 0.4 mg•mL-1 and 80% at 0.8 mg•mL-1 and corresponding times spent were significantly shorter than those of the former three types. There were differences in the varying of reaction rates between type Ⅳ and Ⅴ.【Conclusion】The behavioral characteristics such as reaction rate, reaction time, scavenging rate variation corresponding to reaction process and extract concentration, and EC50 of ethanol extract from sweetpotato sprouting shoot tips duration their DPPH• scavenging reaction should be synthetically taken into account in the assaying antioxidant activity of sweetpotato shoot tips by DPPH• scavenging method.

Transformed GFPuv into Pseudomonas syringae pv. actinidiae and Its Biological Characteristics and Colonization in Soil and Roots of Kiwifruit

HUANG Qi-Ling, GAO Xiao-Ning, ZHAO Zhi-Bo, QIN Hu-Qiang, HUANG Li-Li

Scientia Agricultura Sinica. 2013, 46(2): 282-291. doi:10.3864/j.issn.0578-1752.2013.02.007

Abstract ( 775 )

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【Objective】The objective of this study is to obtain fluorescence-labeled Pseudomonas syringae pv. actinidiae (Psa) strain to further understand the infection and pathogenic processes.【Method】The Psa strain was tagged with GFPuv by electroporation. Fluorescence microscope and the method of plate dilution were used to detect the colonization of tagged strain in soil and root. 【Result】 The transformants showed strong green fluorescence under UV-light indicating GFPuv expression. A 700 bp fragment of the GFPuv gene was amplified from the genome DNA of the transformants. There were no differences between the engineered PSAmx7-GFPuv1 strain and wild-type strain including morphology, optimum temperature, pH and pathogenicity. The green fluorescence could be kept until 20 times of subcultures. The PSAmx7-GFPuv1 could survive in sterile soil for three months, and three weeks or so in non-sterile soil. The PSAmx7-GFPuv1 also could infect and colonise in root.【Conclusion】The GFPuv-tagged Psa was obtained successfully through electroporation in this study. The presence of GFPuv did not have signi?cant impact on the wild-type strain. The tagged strain could survive for a long period of time in sterile soil, colonise and multiply in root.

Fermentation Parameter Optimization of Pythium oligandrum and Biocontrol Effect of the Fermentation Broth

ZHAO Jian, YUAN Ling, HUANG Jian-Guo

Scientia Agricultura Sinica. 2013, 46(2): 292-299. doi:10.3864/j.issn.0578-1752.2013.02.008

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【Objective】 The objective of this study is to optimize the conditions of producing Pythium oligandrum fermentation broth and establish a theoretical foundation for biological control of tobacco black shank. 【Method】 The fermentation parameters were optimized through a single factor and orthogonal experiments, and the biocontrol effects of the fermentation broth were researched by tests on Phytophthora nicotiana, leaves of tobacco in vitro and potted plants in greenhouse. 【Result】 The biomass increased continuously with the culture time, but the inhibition rate increased to the peak then had downward trend. Under the condition of different culture media, temperature, initial pH, illumination time, the more the biomass of mycelia had, the stronger inhibition activity would express, and the correlation coefficients was 0.77**(n=16). Under the condition of culture medium composed of soybean cake powder, initial pH 6.0, 30℃, incubating 6 d and 24 h•d-1 illumination, the biomass of mycelia was the highest and the fermentation broth reached 78.8% for the growth of mycelia and 52.9% for the germination of spore on P. nicotiana. Moreover, the disease incidence and disease index were significantly decreased, the control effect was 69.3%-75.3% in the test on leaves of tobacco in vitro and 55.0%-61.3% in potted plants in greenhouse. 【Conclusion】 In the process of P. oligandrum fermentation, selecting the suitable conditions of the culture medium, pH value, temperature, illumination time and fermentation time, the best inhibition of tobacco black shank can be obtained.

Effects of Long-Term Fertilizer Application on Soil Organic Carbon Change and Fraction in Cumulic Haplustoll of Loess Plateau in China

FAN Ting-Lu, WANG Shu-Ying, ZHOU Guang-Ye, DING Ning-Ping

Scientia Agricultura Sinica. 2013, 46(2): 300-309. doi:10.3864/j.issn.0578-1752.2013.02.009

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【Objective 】The objective of this study was to clarify the effect of long-term fertilization on soil organic carbon (SOC) changes and carbon fraction differences, and to reveal C sequestration and soil fertility improvement model of fertilization, in Cumulic Haplustoll of Loess Plateau in China.【Method】Grain yields and SOC recorded from a 32-year dryland fertilization trial with six fertilizations were used to estimate C input and sequestration and SOC trends, and soil samples in the 32nd year taken from the experiments were separated into sand, coarse silt, fine silt, coarse clay, fine clay, and aggregates by physical way, and the content and distribution of SOC in these particle-size fractions and aggregates were analyzed. 【Result】Cumulative C input ranged from 8.10 t C?hm-2 for the unfertilized treatment (CK) to 69.40 t C?hm-2 for the treatment that received straw (S) plus inorganic N added annually and P fertilizers added every second year (SNP), and there was a difference of 1.82 times in SOC stock in the 0–20 cm soil layer among the fertilizer treatments and SOC stock was 21.76, 24.04, 16.01 t C?hm-2 in the M, MNP, SNP treatments in 2010. SOC stock increased with cumulative C input, C sequestration into SOC rate ranged from 25.80% to 36.05% but 8.20% from straw and root in the SNP, suggesting manure and root residues is more effective in building soil C than straw, SOC significantly built up with time except the CK, in which SOC remained almost stable by root resides C in input only, and annual soil-C sequestration rates (slopes for linear regression of SOC relate to time) were 0.246, 0.326, and 0.361 t C?hm-2?a-1 for the SNP, M, and MNP treatments respectively. SOC in the sand fraction for the MNP and SNP were 3.85 and 2.94 times higher than that for CK, and 2.41 and 1.84 times higher than that for the N. Compared with the CK and NP, SOC in total soil for the MNP increased by 32.50% and 18.10%, but SOC in the sand increased by 285.12% and 105.74%, implying SOC in the sand was the most sensitive fraction to the fertilization among the particle-size organic carbon fractions. However, fertilization increased the ratio of POC (particulate organic carbon) to MOC (mineral-incorporated organic carbon), and the ratio was up to 18.62%, 16.24%, 14.41% for the MNP, M, SNP，9.11% and 9.99% for the N and NP, resulting in improvement of soil active organic carbon and soil fertility. For the CK, SOC in macroaggregates (＞250 µm) was 9.14 times higher than that in microaggregates (＜53 µm), and fertilization increased the SOC by 15.83-23.84 times, and enhanced C/N ratio in macroaggregates, but had less effect on the ratio in microaggregates. SOC in aggregtaes increased with aggregate size, but the effect of organic C increrase in the ＞250 µm aggregates on soil C sequestration rate was higher significanly than that in the ＜53 µm aggregates, implying microaggregate plays an important role in soil organic C sequestration and physical protection. 【Conclusion】Long-term additions of organic materials and inorganic fertilizers, a friendly-environment soil fertility enhancement way for these dryland soils in the China Loess Plateau, significantly promoted C sequestration and sharply resulted in a substantial build-up of SOC due to C inputs from manure and straw and root residues, and the newly organic C was mainly accumulated in the sand and aggregates.

Effect of Long-Term Different Fertilization Management on Weed Community in Rice-Wheat Rotation Field

SHI Lin-Lin, SHEN Ming-Xing, JIANG Min, LU Chang-Ying, WANG Hai-Hou, WU Tong-Dong, ZHOU Xin-Wei, SHEN Xin-Ping

Scientia Agricultura Sinica. 2013, 46(2): 310-316. doi:10.3864/j.issn.0578-1752.2013.02.010

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【Objective】 The characters and effects on weed community caused by different long-term fertilization managements was evaluated in rice-wheat rotation field in Taihu Lake region for the purpose of bio-diversity protection, crop yield increase and noxious weeds decrease. 【Method】 Inorganic fertilizer treatments were selected as follows: CK (no fertilizer), N (N fertilizer), NK (N plus P fertilizer), NPK (N plus P and K fertilizer), NP (N plus P fertilizer) and PK (P plus K fertilizer). Weed density and community surveys were conducted, and bio-diversity indices were also calculated, including diversity index (Shannon-H’), evenness index (Pielou-EJ), and richness index (Margalef-DMG). At the same time, principal component analysis (PCA) and regression analysis were executed to mining the relationship between fertilizer management and weeds.【Result】Total weed density order was NP＞CK＞N＞NK＞NPK, and Lapsana apogonoides, Malachium aquaticum, Beckmannia syzigachne and Vicia sativa were dominant species in Taihu Lake rigion. The biodiversity indices were increased when nitrogen and phosphorus was deficient, and especially, Shannon-H’ index order was CK＞NK＞PK＞N＞NP＞NPK. The eigenvalue of principal components F1 and F2 were 0.498 and 0.235, respectively. L. apogonoides, Geranium carolinianum, Cyperus difformis and B. syzigachne could illustrate more about F1, since the angels were small between F1 and these weeds arrows. The regression coefficients of determination were 0.85, 0.86 (P＜0.05), which analyzed between the value of F1 and soil available-N and Olsen-P, respectively, and there was no significant relationship between F2 and soil nutrient in eigenvalue.【Conclusion】The weed density and dominant species in rice-wheat rotation field were limited by soil nutrient, the biodiversity indices of weed community were higher when soil nutrient was not so sufficient, and weed community was affected sensitively by available-N.

The Impact of Plastic Film Mulching on Citrus Fruit Sugar Accumulation and Sucrose Metabolism-Related Enzymes

JIANG Ni, GAO Hai-Wen, JIN Long-Fei, LIU Yong-Zhong, PENG Shu-Ang

Scientia Agricultura Sinica. 2013, 46(2): 317-324. doi:10.3864/j.issn.0578-1752.2013.02.011

Abstract ( 739 )

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【Objective】 The aim of this research is to investigate the possible physiological mechanism of the effect of plastic film mulching on citrus fruit sugar accumulation, which could help to enrich the knowledge of citrus high fruit quality formation. 【Method】 ‘Egan 1’ Ponkan (Citrus reticulata cv. Egan 1) was used as material, sugar accumulation and activities of sucrose metabolism-related enzymes were compared in fruit segment membrane and juice sacs, as well as contents of leaf total soluble sugar and starch between the plastic film mulching treatments and the control. 【Result】 Plastic film mulching at fruit ripening stage reduced soil water potential, which in the end increased the total soluble sugar content in leaf, fruit segment membrane and juice sacs. On the other hand, the activities of acid invertase and sucrose synthase (cleavage direction) in the segment membrane were higher, while in the juice sac the cleavage direction activity of sucrose synthase was lower in the plastic film mulching fruit than those in the control fruits. 【Conclusion】 Plastic film mulching caused water deficit stress, which induced osmotic adjustment and then to enhance sugar accumulation of fruits. Of them, the changes of sucrose synthase activities in segment membrane and juice sacs under plastic film mulching played an important role in increase of sink strength, thus increasing sucrose input and then enhancing sugar accumulation.

Cloning and Prokaryotic Expression of Flavonoid O-methyltransferase from Camellia sinensis

MA Cheng-Ying, 吕Hai-Peng , LIN Zhi, ZHANG Yue, GUO Li, TAN Jun-Feng

Scientia Agricultura Sinica. 2013, 46(2): 325-333. doi:10.3864/j.issn.0578-1752.2013.02.012

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【Objective】Cloning, sequence analysis of the CsFOMT genes from Camellia sinensis and expression of recombinant proteins in Escherichia coli were conducted to further research the enzymatic characteristics and the metabolic mechanism of regulating the flavonoid in tea plant. 【Method】 The CsFOMT cDNA fragments were amplified from Camellia sinensis by reverse transcription PCR (RT-PCR) and RACE, and then the cloned genes of CsFOMT were inserted into vector pET-28a. The recombinant plasmids pET-28a-CsFOMT were expressed in the prokaryotic expression system after its transformation into E. coli BL21. 【Result】 The whole cDNA sequence was 1 246 bp which contains an ORF of 1 077 bp and encodes 358 amino acids. The putative protein of the gene had an isoelectric point of 5.62 and a calculated molecular weight of 40.2 kD. The coding nucleotide sequence of tea showed 80% and 81% identity with that of Vitis vinifera and Populus trichocarpa, respectively. Results of SDS-PAGE showed that the specific fusion protein was successfully induced to express by IPTG. 【Conclusion】In this study, a full length cDNA of flavonoid O-methyltransferase gene was obtained from Camellia sinensis by RT-PCR and RACE，and the prokaryotic expression vector for this gene was constructed, and then successfully induced to express by IPTG in BL21.

Effects of Ultra-Dry Treatment on Vigor and Antioxidative Metabolism of Pugionium Seeds

ZHAO Peng, ZHANG Yi-Ting, HAO Li-Zhen, PANG Jie, YANG Zhong-Ren, ZHANG Feng-Lan

Scientia Agricultura Sinica. 2013, 46(2): 334-342. doi:10.3864/j.issn.0578-1752.2013.02.013

Abstract ( 631 )

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【Objective】 The effect of ultradry treatment on dynamic change of vigor, antioxidative metabolism of Pugionium. seeds were determined, which would provide a theoretical basis for desiccation tolerance and storage of the seeds. 【Method】The moisture content of P. cornutum(L.) Gaertn. and P. dolabratum Maxim. were reduced to 3.1%, 2.2%, 1.3% from 4.5% and 4.3% (CK), respectively, with silica gel, and the vigor and antioxidative metabolism were detected when the seeds were re-moistened. 【Result】 The vigor of P. dolabratum Maxim. seeds maintained at a relatively higher level, while the P. cornutum(L.) Gaertn. seeds decreased sharply when the moisture content dropped to 1.3% from respective CK. Relative electrical conductivity of the two species seeds with ultradry treatment were lower than the CK and the integrity of cell membranes was conserved. Ultradry treatment accelerated the generation of superoxide generation rate ( ) and malondialdehyde (MDA), but the treatment did not affect the activities of antioxidant enzymes of the two species seeds obviously such as SOD, CAT, GPX, APX. 【Conclusion】The results showed that both of the seeds of P. cornutum(L.) Gaertn. and P. dolabratum Maxim. had strong desiccation tolerance, however, the ability of resisting damage and the desiccation tolerance of P. dolabratum Maxim. seeds were stronger than P. cornutum(L.) Gaertn. seeds, and the moisture content of 2.2% was the turning point of germination and vigor of P. cornutum (L.) Gaertn. seeds.

Structures and Formation Mechanism of Diverse Classes of Dietary Anthocyanin Derivatives

HE Jing-Ren, KUANG Min-Jie, DENG Li, LIU Zhi-Wei, YI Yang, YANG Ning, DUAN Li-Ge

Scientia Agricultura Sinica. 2013, 46(2): 343-355. doi:10.3864/j.issn.0578-1752.2013.02.014

Abstract ( 638 )

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Anthocyanins are one class of natural water-soluble plant pigments which are commonly known to people and possess many important physiological functions and bioactivities. They are important substances responsible for the colours of fruits, vegetables, beans, grains, flowers and medicinal plants. In recent years, anthocyanin derivatives are found in the processed products of fruits and vegetables and some special plants rich in anthocyanins, which are series of natural products derived from anthocyanins and formed during fermentation and ageing or biologically synthesized by metabolization in plants tissues, constituting different families of anthocyanin derivatives with different structures and colours. The present paper summarized the origins of occurrence, structural features and formation mechanism of different types of anthocyanin derivatives (anthocyanin-flavanols, anthocyanin-alkyl-polyphenols, pyranoanthocyanins with the fourth pyran ring linked with different groups, anthocyanidin-like derivatives with flavylium ion and other unique anthocyanin derivatives), which will offer beneficial references for further investigate the structural stability and reactivity of anthocyanins with their applications in processing foods and agricultural products.

Drying Characteristics and Model of Purple Sweet Potato in Air-impingement Jet Dryer

LI Wen-Feng, XIAO Xu-Lin, WANG Wei

Scientia Agricultura Sinica. 2013, 46(2): 356-366. doi:10.3864/j.issn.0578-1752.2013.02.015

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【Objective】In order to improve the drying quality and drying efficiency of purple sweet potato (PSP), the effects of different conditions on PSP air-jet impingement drying characteristics were studied and the optimum drying model was sereened out.【Method】The PSP are dried by air-impingement jet drying. The influence on the drying characteristics and the effective diffusion coefficient under different conditions such as air temperature, air velocity, pre-treatment and slice thickness were all studied. According to statistical calculations, there were 6 drying models were selected.【Result】Compared to the most results of food material drying test, air-impingement jet drying of PSP mainly occurs in the falling rate drying period. Pre-treatment increased the material initial temperature, and led material to an increase of its desired temperature, but reduced the drying rate and extended drying time. The drying rate decreased with the slice thickness increase, while increase with the wind temperature and wind speed increase. The slice thickness and air velocity had a little influence on material temperature increase while the air temperature had a great influence. With the increase of the air temperature value, the time of material reaching the desired environment temperature was increased. The effective diffusion coefficient increased with the increase of film thickness, air temperature and air velocity. The maximum effective moisture diffusion coefficient was 7.0033×10-10 m2•s-1. The drying data were fitted to all the six models. The Modified Henderson and Pabis model gave the highest coefficient of determination and the lowest chi-square and root mean square error. 【Conclusion】 Air temperature, slice thickness, and the pre-treatment have influence on the drying curve, drying rate curve and temperature rise, effective moisture diffusion coefficient. The Modified Henderson and Pabis model could properly describe the air-impingement jet drying behavior of PSP and could be used when the drying air temperatures between 50 and 80℃, air velocities between 10 and 13 m•s-1 and slice thickness between 1.87 and 4.80 mm .

Cloning and Testicular Expression Level of b-Sycp2 in Cattle, Yak and Cattle-Yak

LUO Hua, JIA Chao, QU Xu-Guang, ZHAO Xing-Bo, ZHONG Jin-Cheng, XIE Zhuang, LI Qi-Fa

Scientia Agricultura Sinica. 2013, 46(2): 367-375. doi:10.3864/j.issn.0578-1752.2013.02.016

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【Objective】The study was aimed to clone the Sycp2 gene sequence of cattle, yak and cattle-yak, investigate the gene structures and its tissue-expression patterns and testicular expression levels in cattle, yak and cattle-yak.【Method】 In silico cloning and clone sequencing were applied to acquire the coding region sequences of Sycp2 gene of cattle, yak and yak-cattle, and molecular characterization were analyzed by bioinformatics software. RT-PCR was applied to analyze the tissue-expression patterns, and real-time PCR was employed to examine the expression levels in cattle, yak and cattle-yak testis.【Result】The full length of the coding region sequences of the Sycp2 gene was 4 365 bp, named b-Sycp2. The b-Sycp2 gene in cattle, yak and cattle-yak encoded 1 454 amino acid residues which included some typical domains such as coiled-coil domain. RT-PCR analysis showed that b-Sycp2 gene was expressed only in the testis. Real-time PCR analysis revealed that the mRNA expression level of b-Sycp2 gene in the testis of cattle and yak was remarkably higher than that of in cattle-yak (P＜0.05). 【Conclusion】The b-Sycp2 gene was cloned successfully, and it expressed only in testis, b-Sycp2 mRNA expression level of cattle and yak represented a dramatic higher than that of cattle-yak.

Screening Differentially Expressed Genes of Skin Tissue of Different Flowers Patterns of Hu Sheep

SUN Wei, NI Rong, YIN Jin-Feng, DING Jia-Tong, ZHANG You-Fa, CHEN Ling, WU Wen-Zhong, ZHOU Hong

Scientia Agricultura Sinica. 2013, 46(2): 376-384. doi:10.3864/j.issn.0578-1752.2013.02.017

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【Objective】The molecular genetic mechanism of China’s unique white lambskin breed different flowers patterns of Hu sheep and hair follicle formation were studied.【Method】The article focused on screening differentially expressed genes of new born Hu-sheep’s skin tissue by Agilent gene chip.【Result】 Differential expression analysis revealed that 1067, 2071 and 3879 differentially expressed genes could be screened from 3 groups of large flowers and small flowers, and compared the 3 groups it could find that there were 137 genes in common. Differentially expressed genes classified by GO (Gene Ontology) analysis mainly involved in cell differentiation, proliferation, apoptosis, growth, immune response, ion transport etc. biological progresses. In addition, the RT-PCR results of 3 differentially expressed genes were consistent with that of gene chip.【Conclusion】Combined with the related literatures, BMP7, MMP2, SNAI1, SFXN1, CDKNIC, MT3, POU1F1 etc. might have important effects on the formation of large flowers and small flowers and could be regarded as candidate genes, and offered the basic data for the further study on the molecule genetic mechanism of hair follicles’ formation in Hu sheep.

Transfected FST Gene into Ovine Muscle-Drived Myogenic Cells to Regulate the Expression of MSTN Gene

YUE Qun-Hua, YUAN Jian-Long, HAO Fei, JIN Mu-Zi, WANG Jing-Yuan, YANG Wen-Liang, LIU Dong-Jun, CANG Ming

Scientia Agricultura Sinica. 2013, 46(2): 385-393. doi:10.3864/j.issn.0578-1752.2013.02.018

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【Objective】In this research, skeletal muscle specific expressed follistatin（FST）eukaryotic expression vectors, pCFCDs-red and pSPFCDs-red, were constructed, then respectively transfected into different cell lines. FST and downstream gene expression situation in RNA and protein levels was detected, and the simultaneous start efficiency of skeletal muscle-specific promoter of sub-SP and α-actin was assayed, so that to understand the effect of specific expression of FST on the development of muscle.【Method】Using liposomes, the eukaryotic expression vectors of pCFCDs-red and pSPFCDs-red were transfected respectively into sheep fetal fibroblast cells (SFFCs), skeletal muscle satellite cells (SMSCs) and SMSCs induced muscle tube, and the transgenic cells were analyzed on status of cell growth, cell cycle, cell size and expression of target gene and downstream gene respectively by flow cytometry, Real-Time quantitative PCR and western blotting. 【Result】The growth of transgenic cells basically comply with the trend of non-transgenic cells, and transgenic SMSCs cell had a higher proliferation rates than that genetically modified SFFC. Flow cytometry analysis showed that more than 70% of cells of transgenic cell lines were in G0/G1 phase, maintained a strong ability to divide and had a single peak ,a good aneuploidy and volume increased significantly. Real-time quantitative PCR and western blotting results showed that RNA and protein levels of FST in the transgenic cell lines were raised compared to non-transgenic cell lines. Transgenic SMSCs and myotubes compared to transgenic SFFCs cell had higher expression of FST. Transfected pSPFCDs-red transgenic cells had higher expression level of FST than transfected pCFCDs-red cells.【Conclusion】Skeletal muscle specific promoter SP and α-actin can effectively start FST target gene in the SMSCs and myotubes, and FST have a higher expression level in myotubes. Start efficiency of SP promoter is higher than that of α-actin. In myogenic cells, the increased FST can cause the reduction of MSTN proteins level. FST has a certain role in promoting growth and development of skeletal muscle cells. Key words: FST; synthetic promoter SP ;specific promoter α-actin; skeletal muscle satellite cell; myotubes

Comparative Analysis of Proteome and Phosphoproteome of Drone Embryos Between Apis cerana cerana and A. mellifera ligustica

ZHOU Tian-E, FANG Yu, FENG Mao, HAN Bin, ZHANG Lan, LI Rong-Li, LI Jian-Ke

Scientia Agricultura Sinica. 2013, 46(2): 394-408. doi:10.3864/j.issn.0578-1752.2013.02.019

Abstract ( 637 )

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