华山松大小蠹2个甲羟戊酸路径基因的克隆与表达     

王媛媛  余佳敏  代鲁鲁  陈辉 《西北林学院学报》2020,35(1):140-149

香叶基二磷酸合酶/法尼基焦磷酸（G/FPPS）和异戊二烯焦磷酸异构酶（IDI）是单萜和倍半萜生物合成途径分支点的关键酶。通过克隆华山松大小蠹DaG/FPPS和DaIDI基因的全长序列,预测基因和编码蛋白的性质、结构和功能,并对序列特征和不同发育时期（幼虫、蛹、初羽化期成虫和扩散期成虫）以及不同组织（头、前中肠、后中肠、后肠和剩余躯体）DaG/FPPS和DaIDI表达的研究,旨在揭示DaG/FPPS和DaIDI在华山松大小蠹信息化学物质合成中的分子调控机制。采用RT-PCR和RACE克隆华山松大小蠹DaG/FPPS和DaIDI基因,DNAStar、Blast、ExpasyProtscale、SignalP 4.1、PSORT和TargetP等多种生物信息学方法对基因全长cDNA序列、基本理化性质、系统进化关系、信号肽和蛋白亚细胞定位等进行分析。用Real-time PCR检测DaG/FPPS和DaIDI在华山松大小蠹不同发育时期和不同组织中的表达。结果表明,克隆获得华山松大小蠹香叶基二磷酸合酶/法尼基焦磷酸DaG/FPPS和异戊二烯焦磷酸异构酶DaIDI全长cDNA序列,分别命名为DaG/FPPS和DaIDI,DaG/FPPS编码的氨基酸序列与山松大小蠹DpF/GPPS相似度最高达95%,氨基酸序列含有一个RALS 四肽基序、7 个异戊烯基转移酶保守区（Ⅰ-Ⅶ）和2个典型的DD**D的天冬氨酸富含区;DaIDI编码的氨基酸序列与山松大小蠹IDI相似度最高,达到92%。Expasy Protscale结果表明DaG/FPPS蛋白疏水性氨基酸明显多于亲水性氨基酸,推测其为疏水性蛋白,而DaIDI蛋白亲水性氨基酸数量大于疏水性氨基酸,故推测DaIDI为亲水性蛋白,有较好的水溶性。信号肽预测结果表明,DaG/FPPS和DaIDI均不含信号肽,TargetP结果表明,DaG/FPPS和DaIDI蛋白含线粒体靶向肽,结合PSORT结果推测DaG/FPPS和DaIDI均定位于线粒体。华山松大小蠹DaG/FPPS和DaIDI在不同发育时期和不同组织中均有表达,其中完全羽化期与扩散期成虫保持一致,该阶段表达量最高,不同组织中前中肠表达量最高。  相似文献   

长枝木霉T6菌株对美洲南瓜枯萎病菌的抑制作用     

刘佳  张悦  沈志彦  张树武  薛应钰  梁巧兰  陶飞  徐秉良 《西北农业学报》2020,30(12):1891-1897

为探明长枝木霉T6菌株对美洲南瓜枯萎病菌的作用机理,采用对峙培养、显微观察、孢子萌发、室内防效测定等方法研究其抑制能力和防治效果。结果表明：长枝木霉T6菌株对美洲南瓜枯萎病菌具有较强的抑制作用,对峙培养7 d时,长枝木霉T6菌株对该病菌的抑制率为60.09%;显微形态观察表明：长枝木霉T6菌株菌丝与美洲南瓜枯萎病菌菌丝发生缠绕;长枝木霉T6菌株发酵液对美洲南瓜枯萎病菌分生孢子萌发具有明显的抑制作用,抑制率为31.00%;长枝木霉T6菌株对美洲南瓜枯萎病具有较好的防治效果,室内盆栽防效为69.06%。  相似文献   

加州鲈弹状病毒三水株全基因组序列测定及分析     

刘慧芳  张莉娟  李宁求 《西北农林科技大学学报(自然科学版)》2020,48(3):30-38

【目的】对加州鲈弹状病毒弱毒三水株(MSRV-SS-7)及其母源株(MSRV-SS)进行全基因组克隆和测序分析。【方法】采用分段扩增方法对MSRV-SS-7及MSRV-SS基因组核心序列进行PCR扩增,第1轮根据鳜鱼弹状病毒(SCRV)基因序列(NC_008514.1)设计10对引物进行扩增并测序;在第1轮扩增产物测序、拼接基础上,设计8对引物进行第2轮扩增,拼接后获得基因组核心序列。同时用cDNA末端快速扩增法(RACE)获得3′和5′末端序列。采用Vector NTI 8.0对基因组核心序列和末端序列进行拼接,获得基因组全序列,使用Clone Manager 8.0对MSRV-SS-7和MSRV-SS进行序列比对分析;同时使用DNAMAN将基因组G基因序列转换为氨基酸序列,并通过MEGA 5.0与其他鱼类弹状病毒的G蛋白氨基酸序列进行进化树分析。【结果】MSRV-SS-7及其母源株MSRV-SS基因组全长均为11 548 bp,包含N、P、M、G、L等5个结构基因,基因组结构为3′Leader-N-P-M-G-L-Trailer 5′;MSRV-SS-7与MSRV-SS全基因组中共有10处核苷酸不同。G蛋白进化分析表明,MSRV-SS-7及母源株MSRV-SS与SCRV亲缘关系最近,且与鲈鱼弹状病毒属(Perhabdovirus)聚为一支。【结论】克隆分析了MSRV-SS-7的全基因组序列,可以用于后续活疫苗的开发。  相似文献   

DTA-6对两种食用豆生理代谢及产量的影响   总被引：1，自引：0，他引：1  

丁凯鑫  单莹  冯乃杰  郑殿峰  梁喜龙  吴琼  黄文婷 《作物杂志》2020,36(5):148-50

为探究2-N,N-二乙氨基乙基己酸酯（diethyl aminoethyl hexanoate,DTA-6）对食用豆叶片的生理代谢及产量的调控效应,选用芸豆（英国红）和小豆（龙垦2号）为试验材料,采用大田完全随机试验方法,于芸豆和小豆的初花期叶面喷施DTA-6,以喷施清水为对照（CK）,测定各生育时期叶片光合参数、碳代谢产物、干物质、产量及产量构成因素。结果表明：DTA-6提高了2种食用豆各生育期的SPAD值、盛花期和鼓粒期的净光合速率（P_n）、蒸腾速率（T_r）、气孔导度（G_s）和胞间CO₂浓度（C_i）,显著增加了叶片蔗糖和可溶性糖的含量,促进了叶片淀粉的积累;与CK相比,DTA-6提高了2种食用豆的地上部单株干物质积累量,并且显著提高了芸豆和小豆鼓粒期荚分配率;DTA-6可有效调控2种食用豆的单株荚数、单株粒数和百粒重,从而提高产量,DTA-6处理的芸豆和小豆产量分别较CK增加13.30%和12.91%,增产显著。  相似文献   

Generation of a uniform thymic malignant lymphoma model with C57BL/6J p53 gene deficient mice     

Susu Liu  Jianjun Lyu  Qianqian Li  Xi Wu  Yanwei Yang  Guitao Huo  Qingfen Zhu  Ming Guo  Yuelei Shen  Sanlong Wang  Changfa Fan 《Journal of toxicologic pathology》2022,35(1):25

Lymphoma is the third most common cancer diagnosed in children, and T-cell lymphoma has the worst prognosis based on clinical observations. To date, a lymphoma model with uniform penetrance has not yet been developed. In this study, we generated a p53 deficient mouse model by targeting embryonic stem cells derived from a C57BL/6J mouse strain. Homozygous p53 deficient mice exhibited a higher rate of spontaneous tumorigenesis, with a high spontaneous occurrence rate (93.3%) of malignant lymphoma. Because tumor models with high phenotypic consistency are currently needed, we generated a lymphoma model by a single intraperitoneal injection of 37.5 or 75 mg/kg N-methyl-N-nitrosourea to p53 deficient mice. Lymphoma and retinal degeneration occurred in 100% of p53^+/− mice administered with higher concentrations of N-methyl-N-nitrosourea, a much greater response than those of previously reported models. The main anatomic sites of lymphoma were the thymus, spleen, bone marrow, and lymph nodes. Both induced and spontaneous lymphomas in the thymus and spleen stained positive for CD3 antigen, and flow cytometry detected positive CD4 and/or CD8 cells. Based on our observations and previous data, we hypothesize that mice with a B6 background are prone to lymphomagenesis.  相似文献   

SHK-6对不同群体下大豆花荚脱落及其产量的调控   总被引：6，自引：1，他引：5  

张海峰  张明才  翟志席  段留生  李召虎  黄淮  陈刚 《大豆科学》2007,26(1):78-83

以"冀豆12"为材料,在大田条件下通过在初花期对不同群体大豆施用SHK-6,研究了植物生长调节剂SHK-6对不同群体条件下大豆产量及其构成因素的调控作用.结果表明:SHK-6能够明显的提高各个群体下的大豆产量(8.0%),在密度水平为30株/m2的条件下施用SHK-6产量达到最高.调节剂SHK一6能够提高单株花数(13.0%)、荚数(14.8%)、粒数(10.9%)、荚重(16.4%)、粒重(12.3%),并且能够提高百粒重(2.4%)、粒茎比(15.0%).虽然SHK-6处理也增加不同密度水平下的花荚脱落数,但对脱落率没有影响.SHK-6能够降低叶面积指数(LAI)、提高下层叶片的光截获率(群体透光率)和下层叶片的叶绿素的含量.可见冠层结构的优化是导致产量提高的根本原因.  相似文献   

The effects of various plant protection methods on the development of Zymoseptoria tritici and Cephalosporium gramineum,grain yield and protein profile     

Urszula Wachowska  Iwona Konopka  Adrian Duba  Klaudia Goriewa 《国际虫害防治杂志》2019,65(2):105-113

Septoria leaf blotch progresses rapidly, leading to the development of Zymoseptoria titici forms resistant to fungicides. Cephalosporium stripe is caused by Cephalosporium gramineum. The aim of this study was to evaluate the effectiveness of selected pesticides in limiting the symptoms of both diseases on winter wheat leaves, and to determine their influence on grain yield and the content and composition of protein fractions in wheat kernels. Propiconazoles were most effective in inhibiting the development of Septoria leaf blotch (symptoms were reduced from 54.7% to 78.6%). Strobilurins were less effective due to the presence of isolates with the G143A mutation. Symptoms of Cephalosporium stripe were rarely observed, and protective treatments did not reduce their severity. The highest content of grain protein (14.81%) was found in plants most intensely protected with the fungicides containing fenpropimorph, pyraclostrobin and epoxiconazole. The principal component analysis revealed that the plant protection method influenced the grain protein profile. The accumulation of HMW glutenins and α/β gliadins was mutually interrelated and higher in high-input treatments; control grain was characterized by close relationships between ω-gliadins, LMW glutenins, albumins and globulins, whereas low-input treatments influenced mostly γ-gliadins.  相似文献   

草鱼呼肠孤病毒VP6蛋白在毕赤酵母中表达的初步研究   总被引：1，自引：0，他引：1  

苏岚  曾令兵  周勇  张辉  高正勇  张金凤 《淡水渔业》2012,42(6):38-42

根据GeneBank中草鱼呼肠孤病毒(grass carp reovirus,GCRV)104株VP6蛋白的全基因序列(HM234682)设计一对特异引物,以提取的GCRV-104核酸为模板,采用RT-PCR技术扩增VP6蛋白编码基因,并将其克隆到含强启动子AOXⅠ的毕赤酵母(Pichia pastoris)表达载体pPICZB上。重组酵母质粒pPICZB-VP6经SacⅠ线性化,电击转化到毕赤酵母KM71菌株中,Zeocin抗性平板上筛选高拷贝转化子。阳性转化子在30℃,0.5%(V/V)的甲醇添加量的条件下,连续诱导3 d,表达产物经SDS-PAGE和Western-Blotting检测,结果表明成功实现了GCRV-104 VP6蛋白在毕赤酵母中的胞内表达,重组VP6蛋白相对分子质量约46 000,与天然VP6蛋白相对分子质量接近,并且可与鼠抗草鱼GCRV-104 VP6蛋白的多克隆抗体特异性结合。  相似文献   

中华绒螯蟹FAD6-b基因的全长克隆及原核表达分析     

杨志刚  姚琴琴  成永旭  施秋燕  杨青  何杰  马明君  王瑶  常东 《水产学报》2016,40(1):24-35

根据已登录的中华绒螯蟹一种Δ6去饱和酶基因(Accession Number:JX946434)及菁夜蛾去饱和酶基因(Accession Number:KJ622055.1)的保守区设计引物,通过逆转录PCR以及RACE技术克隆了中华绒螯蟹另一种Δ6去饱和酶基因FAD6-b的全长序列。经序列分析,中华绒螯蟹FAD6-b cDNA全长为2 310 bp,其中开放阅读框(ORF)为1 326 bp,共编码442个氨基酸(Accession Number:KP876058),理论分子量为50.86 ku,理论等电点为8.47,FAD6-b基因的氨基酸序列与已公布的一条中华绒螯蟹Δ6去饱和酶基因的一致性为76%。原核表达载体构建及其表达实验表明,中华绒螯蟹FAD6-b基因成功重组到原核表达载体pCold-TF DNA中,重组质粒pCold TF-fad6b在大肠杆菌BL21(DE3)pLysS中进行表达,经SDS-PAGE电泳分析表明,IPTG诱导后的重组菌出现了单一的蛋白条带,且大小与预期(105.86 ku)一致,可溶性分析显示目的蛋白主要存在于蛋白上清液中。对重组蛋白进行纯化,蛋白纯化液经电泳检测后显示出特异性单一条带,进一步证明了pCold TF-fad6b的成功构建和表达。目的蛋白经Western-blotting检测,结果表明获得的重组蛋白与6×His抗体进行了特异性结合,显示出良好的免疫学活性。  相似文献   

Structural and Rheological Characteristics of Alginate from Sargassum cristaefolium Extracted by Twin Screw Extruder     

Sugiono Sugiono  Masruri Masruri  Simon Bambang Widjarnako 《Journal Of Aquatic Food Product Technology》2013,22(9):944-959

This study aimed to investigate the effects of process extrusion on the characteristics of Sargassum cristaefolium sodium alginate (SCSA) extracted using twin-screw extruder. Box-Behnken Design (BBD) from response surface methodology (RSM) was established to understand the effects of temperature, screw speed, and pH on the multiple-responses of alginate characteristics including intrinsic viscosity, yield, and molecular weight. The results revealed that temperature, screw speed, and pH significantly affected (P < .05) all responses. The optimum extraction condition was found at temperature of 58.18°C, screw speed of 77.99 rpm, and pH 10.11. At this condition, the response of residence time distribution was 7.07 ± 0.029 min, yield of 34.01 ± 0.12%, intrinsic viscosity of 460.13 ± 14.75 mL/g, and molecular weight of 217.94 ± 7.14 × 10³ g/mol. This alginate had mannuronic acid to guluronic acid (M/G) ratio of 0.28, and the L-guluronic acid block was 0.78, which was higher than the D-mannuronic acid block. Rheological characterization of SCSA in aqueous solution was shear-thinning pseudoplastic, and alginate gel in 1 M CaCl₂ was more elastic than liquid.  相似文献