Genome-Wide Analysis of the F3′5′H Gene Family in Blueberry(Vaccinium corymbosum L.)Provides Insights into the Regulation of Anthocyanin Biosynthesis

The F3′5′H gene family plays an important role in regulating anthocyanin biosynthesis,abiotic stress,and hormone signaling.In this study,14 F3′5′H genes were identified from the blueberry genome.The chromosomal distribution,physicochemical properties,F3′5′H domain,conserved motifs,cis-acting elements,and intron/exon compositions were analyzed.The functional prediction analysis of these VcF3′5′Hs indicated that their biological functions included light response and other secondary metabolites.The results of qRT-PCR showed that VcF3′5′Hs(especially VcF3′5′H4)were highly expressed at the ripening stage.Subcellular localization revealed that VcF3′5′H4 may be located in the endoplasmic reticulum.Co-expression analysis showed that the VcF3′5′H gene family was related to anthocyanin.This research provides an overview of the blueberry F3′5′H family and helps verify the role of these genes in regulating anthocyanin biosynthesis.

H型高血压急性缺血性脑卒中患者亚甲基四氢叶酸还原酶C677T基因多态性及其与肾功能的相关性

目的观察H型高血压急性缺血性脑卒中患者亚甲基四氢叶酸还原酶(MTHFR)C677T的基因多态性,分析其与H型高血压急性缺血性脑卒中患者肾功能的相关性。方法选择153例H型高血压急性缺血性脑卒中患者为观察组,同期158例非H型高血压急性缺血性脑卒中患者为对照组。两组均采集外周静脉血,采用PCR扩增和微阵列技术检测MTHFR C677T基因型,测算全身免疫炎症指数(SII),采用日立7600型全自动生化分析仪检测两组血清肌酐,据此测算肾小球滤过率(eGFR)。采用多元线性回归分析法分析MTHFR C677T基因型与H型高血压急性缺血性脑卒中患者同型半胱氨酸(Hcy)、eGFR的相关性,采用Spearman相关分析法分析SII与H型高血压急性缺血性脑卒中患者eGFR、Hcy的相关性。结果与对照组相比,观察组患者TT基因型分布频率最高,T等位基因频率最高(χ^(2)分别为19.188、5.138,P均<0.05)。观察组、对照组患者SII分别为583.54(384.97,903.73)、425.03(310.26,583.16),二者相比,P<0.05。与CC、CT基因型比较,TT基因型的H型高血压急性缺血性脑卒中患者血清Hcy水平高,eGFR水平低(F分别为28.544、3.749,P均<0.05)。MTHFR C677T TT基因型与H型高血压急性缺血性脑卒中患者血清Hcy呈正相关(β=4.173,P<0.05),与eGFR呈负相关(β=-6.559,P<0.05)。SII与H型高血压急性缺血性脑卒中血清Hcy水平呈正相关(r=0.226,P<0.05),与eGFR呈负相关(r=-0.129,P<0.05)。结论H型高血压急性缺血性脑卒中患者MTHFR C677T基因型主要为TT型。MTHFR C677T TT基因型的H型高血压急性缺血性脑卒中患者可能更易引起肾功能下降。

Association of Host Interferon-γ Gene Polymorphism with Toxoplasma gondii Infection in Pregnant Women of Bangladesh

Human toxoplasmosis is caused by the intracellular protozoan parasite Toxoplasma gondii. Although T. gondii infection is generally asymptomatic for most of the immunocompetent adults, severe complications may occur particularly in pregnant women and immunocompromised individual. Host cell immunity plays a critical role in parasite differentiation and persistence in the host. Therefore, genetic polymorphism in the host immune genes, for instance interferon-γ gene could be linked with possibility of T. gondii infection. The objective of the study was to verify the link between the single nucleotide polymorphisms (SNPs) in the IFN-γ gene of pregnant women and T. gondii infection through correlating with anthropometric and sociodemographic parameters. In this study, ninety-two (N = 92) pregnant women (16 - 40 years) and healthy controls (N = 95) with similar age ranges were included. Among them, 25% (n = 23) pregnant women were seropositive for T. gondii IgG antibodies by Rapid Test Assay. Allelic and genotypic frequencies of IFN-γ +874T/A (rs2430561) SNPs were evaluated by using ARMS-PCR. The distribution of the A and T alleles in the specific position of the IFN-γ gene in the T. gondii-infected pregnant women and the control groups did not differ significantly, according to the data. However, we found a higher frequency (13.04%) of A/A genotype in T. gondii infected pregnant women as compared to non-infected individuals (8.70%), demonstrating that T. gondii infection susceptibility may be increased by homozygosity for the A allele. Further studies are to be needed to find out the link between host gene polymorphism and T. gondii infection in Bangladesh.

新疆哈萨克族H型高血压与MTHFR基因多态性及叶酸水平的相关性研究

目的:探讨哈萨克族H型高血压与同型半胱氨酸(Hcy)代谢相关基因MTHFR多态性和叶酸的关联,为H型高血压发病机制提供理论依据。方法:应用新疆哈萨克族H型高血压现况调查数据库,采用病例-对照研究,随机选取H型高血压患者100例、单纯高血压患者100例、高Hcy血症患者100例,健康人群100例作为对照组。应用Sanger测序法对所有研究对象的MTHFR基因rs1801133、rs1801131位点进行基因型检测。叶酸采用电化学发光法进行检测。采用χ^(2)检验进行Hardy-Weinberg平衡检验;应用二分类非条件Logistic回归分析H型高血压与MTHFR基因位点以及叶酸之间的关联。结果:以健康组为对照,高水平叶酸是哈萨克族H型高血压的保护因素(OR=0.33,95%CI:0.15-0.72),rs1801133-(CT+TT)基因型是危险因素(OR=6.37,95%CI:2.97-13.68);高水平叶酸是哈萨克族高Hcy血症的保护因素(OR=0.45,95%CI:0.23-0.86),rs1801133-(CT+TT)基因型是危险因素(OR=2.92,95%CI:1.52-5.62)。结论:MTHFR基因的rs1801133-T等位基因和rs1801133-(CT+TT)基因型是H型高血压的危险因素;高水平叶酸是H型高血压的保护因素。

茎莴苣F6′H家族基因鉴定及其与鲜切莴苣褐变的关系初探

为深入探究阿魏酰-CoA-6′-羟化酶(feruloyl-CoA 6′-hydroxylase, F6′H)在鲜切莴苣褐变过程中的作用及机制,本研究通过Blastp结合结构域筛选的方法从茎莴苣基因组中搜索鉴定F6′H家族成员,并对其进行生物信息学分析及褐变过程中的表达分析,同时测定了鲜切莴苣贮藏过程中褐变度和总酚含量的变化。结果表明,共鉴定出11个茎莴苣F6′Hs基因,其编码蛋白均为定位于细胞质中的亲水蛋白,氨基酸数在181~840个之间,分子量在20.74~93.35 kDa之间,等电点在5.16~8.36之间,脂肪系数在82.28~102.98之间,不稳定指数为26.13~55.39。鲜切莴苣贮藏过程中LsF6′H9基因明显上调表达,LsF6′H3和LsF6′H11在贮藏4~6 d上调表达,其余基因下调表达或变化较小;褐变度逐渐升高,总酚含量先升高后降低,贮藏6 d时最高。相关性分析结果显示,总酚含量与褐变度呈显著正相关,表明酚类的积累可能是导致鲜切莴苣发生褐变的重要因素;褐变度及总酚含量均与LsF6′H1、LsF6′H2、LsF6′H3、LsF6′H4、LsF6′H9、LsF6′H10和LsF6′H11的表达量呈正相关,特别是总酚含量与LsF6′H9表达量呈极显著正相关,褐变度与LsF6′H9表达量呈显著正相关,表明LsF6′H9可能在鲜切莴苣褐变中扮演重要角色。本研究结果可为深入探究F6′H在鲜切莴苣褐变过程中的作用及机制提供理论依据。

基于H&E图像和基因表达数据的多模态深度学习模型预测胃癌生存风险

胃癌作为高发恶性肿瘤,其致死率近年来居高不下,因此精准预测胃癌患者的生存风险对于治疗至关重要。文章提出了一种基于多模态深度学习的预测模型,旨在评估胃癌患者的生存风险。该模型整合了H&E(Hematoxylin-Eosin staining)染色图像和基因表达数据,首先,采用ResNet18卷积神经网络模型提取深层H&E图像信息,将其编码为一维特征向量。其次,采用多模态紧凑型双线性池化方法,将图像特征与基因表达数据进行融合,用于预测胃癌患者的风险分数。在TCGA的胃癌样本中,该模型的一致性指数(c-index)为0.70。在测试集上进行的Kaplan-Meier分析结果显示,模型成功地区分出高风险群和低风险群。结果表明,该模型在区分胃癌患者风险层次方面表现出色,具有显著优势。

拟南芥TRX-h5基因的克隆和表达分析

硫氧还蛋白(TRX-h)是一类催化二硫键还原的小分子蛋白,在调控氧化应激响应中发挥重要作用。为了研究硫氧还蛋白的结构及其功能,文章以野生型拟南芥(Arabidopsis thaliana)Columbia(Col-0)为研究材料,通过反转录-聚合酶链式反应(reverse transcription-polymerase chain reaction,RT-PCR)技术获得TRX-h5基因的编码序列(coding sequence,CDS),体外构建pET28a(+)-TRX-h5及pET28a(+)-TRX-h5M(第39位和第42位半胱氨酸位点突变)的原核表达载体,转化至大肠杆菌感受态细胞BL21(DE3)中,诱导后经镍柱纯化得到带有6个His标签的融合蛋白。通过SDS-PAGE电泳技术检测到14 kDa处的目的条带,与理论值预测一致。运用生物信息学分析得出,TRX-h5基因编码118个氨基酸,相对分子质量为13122.32,理论等电点为5.19,该蛋白的不稳定参数为26.74,属于稳定性蛋白。点突变分析发现,TRX-h5的第39位和第42位半胱氨酸位点决定了该蛋白的催化活性,为进一步探究TRX-h5的蛋白功能和高等植物氧化还原系统的研究提供了新思路。

HBA2:c.2T>C和HBA2:c.2delT两例罕见突变引起血红蛋白H病家系分析

目的:分别对HBA2:c.2T>C和HBA2:c.2delT两种罕见HBA2基因起始密码子突变复合东南亚型α-地贫的血红蛋白H病病例及其家系成员进行致病基因分析,了解HBA2:c.2T>C和HBA2:c.2delT突变与临床表型的关系。方法:采集家系成员外周血进行血细胞分析及毛细管电泳血红蛋白分析,缺口PCR(Gap-PCR)、反向点杂交法(RDB)检测ɑ-地贫基因常见类型突变,Sanger测序法对HBA1和HBA2基因序列进行分析。结果:检测出两个先证者基因型分别为--SEA/αα复合HBA2:c.2T>C和--SEA/αα复合HBA2:c.2delT,家系成员中检出HBA2:c.2T>C/WT和HBA2:c.2delT/WT,均表现为小细胞低色素性贫血。结论:HBA2:c.2T>C和HBA2:c.2delT为杂合突变时机体可出现静止型α-地贫的表型,当其复合轻型α-地贫时可使机体出现血红蛋白H病的临床表现,本研究为遗传咨询提供依据。

Cloning, Characterization and Chromosome Localization of Two Powdery Mildew Resistance-Related Gene Sequences from Wheat

Reverse_transcription Polymerase Chain Reaction (RT_PCR) was performed using cDNAs as templates from wheat_ Haynaldia villosa 6VS/6AL translocation line and 'Yangmai 5' induced with fungus Erysiphe graminis , and degenerate primers designed based on the conserved amino acid sequences of known plant disease_resistance genes. The cDNA sequences encoding cyclophilin_like and H +_ATPase_like genes were first isolated and characterized in wheat. The putative amino acid sequences of the two clones showed that they were highly homologous to those of cyclophilin proteins and H +_ATPases isolated from other plants. Thus they were designated as Ta_Cyp and Ta_MAH . The obvious expression differences could be observed between wheat_ H. villosa 6VS/6AL translocation line and susceptible wheat cultivar 'Yangmai 5', implying that the two genes may be related with the resistance of wheat_ H. villosa 6VS/6AL translocation line to disease. Southern blot indicated that the wheat genome contained 2-3 copies of Ta_Cyp gene and one copy of the Ta_MAH gene. Chinese Spring nulli_tetrasomic line analysis located the Ta_Cyp homologous genes on wheat chromosome 6A, 6B and 6D. Southern blot using Ta_Cyp clone as a probe showed that the polymorphic bands existed among the H. villosa , amphiploid of Triticum durum _ H. villosa , wheat_ H. villosa 6VS/6AL translocation line and 'Yangmai 5', suggesting that Ta_Cyp homologies exist in wheat genome as well as on the short arm of chromosome 6V in H. villosa .

Construction of Recombinant Pseudorabies Virus Expressing Canine Distemper Virus H Gene and Analysis on Its Biological Characters

[Objective] The aim was to construct a recombinant pseudorabies virus expressing canine distemper virus H gene and investigate its biological characters.[Method] H gene of canine distemper virus（CDV）strain Onderstepoort was produced by RT-PCR,inserted into pcDNA3.1（＋）vector to construct a expression cassette,which was then subcloned into transfer vector p8AA,prior to the insertion of LacZ expression cassette.The resulting new transfer vector was named as p8AAZH.Subsequently,p8AAZH was co-transfected with the genome of pseudorabies virus（PRV）Bartha-K61 into BHK-21 cells to enable gene recombination and virus package,and the virus solution was collected as cytopathic effect occurring.A series of procedures including blue plaque purification,PCR identification,observation under electron microscope and Western blot were carried out to screen the recombinant pseudorabies virus and identify the protein expression of target gene.Meanwhile,growth curve of the recombinant virus was determined in BHK-21 cells.[Result] The H gene had been inserted into the genome of Bartha-K61 strain,and RPRV-H was the same as Bartha-K61 in the one-step growth curve and cytopathic effect in BHK-21 cells.[Conclusion] The recombinant pseudorabies virus was constructed,and the insertion of H gene did not influence proliferation of recombinant virus,which laid a foundation for development of recombinant canine distemper virus vaccine.

Construction and Identification of a Goat Pox Virus Transfer Vector to Express Peste des Petits Ruminants H gene

[Objective] This study was to develop a live vector vaccine of goat pox virus of Peste des petits ruminants(PPR). [Method] Using PCR amplification technique, PPR H gene was obtained, then ligated into pGEM-T easy vector; the recombinants were digested by Nhe Ⅰ and Hind Ⅲ, and ligated into pEGFP-N1-P7.5, yielding the recombinant vector pEGFP-N1-P7.5-H; next the expression cassette EGFP-N1-P7.5-H was first released from recombinant vector pEGFP-N1-P7.5-H by double digestion of Hind Ⅲ and Nhe Ⅰ and ligated into pUC119-TK that was digested by Kpn Ⅰ, yielding the transfer vector pUC119-TK-EGFP-P7.5-H. [Result] Identification and double enzyme digestion showed that the transfer vector pUC119-TK-EGFP-P7.5-H was correctly constructed. From the transfer vector transfected BHK-21 cells which infected GTPV AV41, specific fluorescence was observed at 48th h of transfection. [Conclusion] The construction of goat poxvirus live vector laid a foundation for the live vector vaccine of PPR vaccine.

Screening an Na^+/H^+ Antiporter Gene from the Halophiles Colonizing in the Dagong Ancient Brine Well of Zigong City,China

[Objective] This study aimed to screen an Na＋/H＋ antiporter gene from the halophiles colonizing in the Dagong Ancient Brine Well in Zigong City, China, and then analyze the gene structure and properties of the protein encoded by this gene. [Method] Metagenomic DNA libraries of halophiles from the Dagong Ancient Brine Well were used for screening genes with Na＋/H＋ antiporter activity in antiporter-defi- cient E. coil KNabc strain by functional complementation. Then the start codon, stop codon, ORF, -35 region, -10 region and SD sequence of Na~/H＋ antiporter gene, as well as the molecular weight, isoelectric point, hydrophobic region, transmembrane domain, phyletic evolution and salt resistance of protein encoded by the gene were investigated. [Result] A new Na＋/H＋ antiporter gene m-nha was obtained, which ,ren- dered the antiporter-negative mutant E. coil KNabc cells with both the resistance to Na＋ and the ability to grow under alkaline conditions. [Conclusion] The structure and amino acid sequence of M-Nha was different from the previously reported Na＋/H~ antiporters, and the m-nha gene disclosed from the Dagong Ancient Brine Well was identified as a novel Na＋/H＋ antiporter gene. This study was significant not only in helping us understand the salt tolerance of halophiles in ancient brine wells and develop and utilize the genes resource, but also in exploring new salt-tolerant genes.

Sequence Analysis of HA Genes from Three H9N2 Subtype Avian Influenza Viruses

[ Objective] The study aimed to understand the genetic characters of H9N2 subtype avian influenza viruses isolated in Belling area. [ Method] HA genes of three H9N2 subtype avian influenza viruses A/Chicken/Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/ liu/00 were amplified by RT-PCR and then sequenced. [ Result] The results of phylogenetic analysis showed that A/Chicken/Beijing/xu/00, A/ Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 shared the nucleotide homologies of 84.8% （ Dk/HK/Y439/97 ） -98.0% （ Ck/GX17/00 ）, 85.1% （Dk/HK/Y439/97） - 99.1% （ Ck/GXl 7/00）, 90.7% （ Ck/BJ/3/01 ） - 99.1% （Ck/GX17/00） with the isolates from Hongkong and other are- as of Chinese Mainland respectively. At the same time, the analysis of amino acid indicated that the three isolates belonged to low pathogenic H9N2 isolates of avian origin. The 226^th amino acid of them were L （ Leu）, suggesting their high binding affinity to human cells. There were seven glyco- sylation sites in HA protein, five from HA1 and two from HA2. [ Cenclusien] By analysis at molecular level, it could be concluded that A/Chicken/ Beijing/xu/00, A/Chicken/Beijing/bei/00 and A/Chicken/Beijing/liu/00 were low pathogenic H9N2 isolates of avian origin.

Sequence Comparison and Analysis of HA Gene of Four H9N2 Avian Influenza Virus Isolates

[ Objective] To determine the HA gene sequences of four H9N2 Avian influenza virus （AIV） strains and carry out comparative analysis so as to understand the difference and variation pattern of each strain from the angle of molecular biology and to know the distribution and epidemic law of H9N2 AIV. [Method] One pair of primers was designed referring to HA gene sequences of H9N2 AIV. The HA genes of A/Chicken/Hebei/WD/98 （H9N2; WD98 for short）, A/Chicken/Hebei/ZD/04 （H9N2; ZD04 for short））, A/Chicken/Beijing/MY/06 （H9N2; MY06 for short） ）, and A/Chicken/Beijing/PG/08 （H9N2; PG08 for short）） were amplified, cloned and sequenced. Then the HA gene sequences of these strains were compared with that of 10 H9N2 AIV stains in GenBank. [Result] The ORF of HA genes of the four strains was 1 683 bp in size, encoding 516 amino acids. The HA gene sequences of the four strains, WD98, MY06, PG08, and ZD04, were 82.6% -95.1%, 83.0% -99.0%, 82.7% -95.5%, and 81.3% -95.7% homologous to that of the 10 H9N2 AIV stains, respectively. And the homology of amino acid was respectively 86.6% -96.3%, 86.6% -97.9%, 87.0% -97.1%, and 86.9% -97.3%. [ Conclusion] The HA gene has greatly high homology among different strains.

Cloning and Phylogenetic Analysis of NS1 Genes from Different Isolates of H9N2 Subtype Duck Influenza Virus

[ Objective] The study aimed to lay a foundation for the further studies on function mechanism of NS1 protein in the interspecies transmission of waterfowl influenza virus. [Method] Using the serologic assay and the specific RT-PCR method, some strains of H9 subtype waterfowl influenza virus were isolated from the 12 to 20 day-old muscovy duck flocks without any clinical symptoms in different areas of Guangdong Province. Four of these strains, including A/duck/ZQ/303/2007（H9N2） （A3 for short）, A/Duck/FJ/301/2007 （H9N2） （C1 for short）, A/Duck/NH/306/2007（H9N2） （ D6 for short）, A/duck/SS/402/2007（H9N2） （ E2 for short）, and a strain named A/duck/ZC/2007（H9N2） （L1 for short） from a muscovy duck died of avian influenza virus （AIV）, were used for NSl gene cloning and sequencing. Subsequently, the obtained NSl gene sequences were compared with other NS1 sequences registered in GenBank, and the phylogenetic analysis was also conducted. [Result] When compared with the H9N2 AIV NS1 sequences in GenBank, the NSl genes of the four AIV strains A3, C1, 136 and E2 displayed homologies ranging from 99% to 100% at nucleotide level, and 95% to 100% at amino acid level; while the NSl gene of L1 strain displayed homology ranging from 94% to 97% at nucleotide level, and 93% to 98% at amino acid level. The phylogenetic tree demonstrated that A3, C1, D6 and E2 were highly resemblant, and L1 was closest to AY66473 （chicken, 2003）. By comparison with the NS1 gene sequences of L1, AF523514 （duck）, AY664743 （chicken） and EF155262.1 （quail） using DNAstar, A3, C1, D6 and E.2 presented nucleotide variations at site 21 （ R→Q）, 70, 71 （ KE→EG）, 86 （ A→S）, 124 （V→M） and 225 （ S→N）, and amino acid variations at site 21,70, 71 and 86 in dsRNA- dependent protein kinase （PKR） binding domain of NSl gene, which induced the evident variations of antigenic determinant and surface proba- bility plot of NS1 protein. [ Conclusion] This study suggested that the amino acid sequence variation in PKR binding domain of NS1 protein had something to do with the virus pathogenicity.

Sequence Analysis of Type III Effector tccP and tccP2 Genes in Escherichia coli O157:H7 from Chinese Water-chestnut

[Objective] This study aimed to analyze the type III effector tccP and tccP2 genes in Escherichia coli O157：H7 from Chinese water-chestnut. [Method] Gene-specific and locus-specific primers were utilized to amplify tccP/tccP2 and their flanking regions for sequence analysis. [Result] E. coli O157：H7 CWN11 harbored intact tccP and tccP2 genes, however, the number of proline-rich repeats in tccP gene was only one that probably resulted in biological incapability, whereas, the tccP2 gene consisted of five and half proline-rich repeats and could encode functional protein. [Conclusion] Here, we reported the first sequence of tccP gene that consisted of only one proline-rich repeat and tccP2 was assumed to play a crucial role in colonization and subsequent signaling cascades.

犬瘟热病毒N和H蛋白拮抗IFN-β信号通路的研究

为研究犬瘟热病毒(Canine distemper virus, CDV)不同毒株N、H蛋白调控宿主β-干扰素(IFN-β)表达的作用,将CDV野毒株LN(10)1、SD(14)7和疫苗株CDV3感染提前转染了pGL3-IFN-β报告基因质粒和pRL-TK内参质粒的MDCK细胞,分别在感染0 h、8 h、12 h和24 h后通过双荧光素酶报告系统检测试剂盒测定IFN-β启动子的活性。将CDV野毒株LN(10)1、SD(14)7和疫苗株CDV3 N和H基因克隆到pCI真核表达载体。重组表达质粒分别转染HEK293T细胞,应用Western blot检测蛋白能否正确表达。将N和H蛋白重组表达质粒连同pGL3-IFN-β报告基因质粒和pRL-TK内参质粒共转染至HEK293T细胞后,经Poly(I:C)刺激细胞,利用双荧光素酶报告系统检测系统测定IFN-β启动子活性。结果显示3种不同毒力的CDV在感染MDCK细胞后,只有CDV 3疫苗株感染后能观察到IFN-β启动子短暂激活。Western blot实验证实,3种毒株N和H蛋白在HEK293T细胞中均能正确表达。双荧光素酶报告系统对IFN-β启动子检测活性结果显示,Poly(I:C)成功激活了IFN-β信号通路,而这种激活作用在表达CDV不同毒株的N或H蛋白时均受到显著抑制(P <0.01),但不同毒株N或H蛋白对IFN-β信号通路的调控与CDV毒力无关。本研究证实了CDV野毒株和疫苗株N、H蛋白可抑制宿主IFN-β信号通路,为进一步研究CDV与宿主分子互作奠定基础。

怀化地区犬瘟热病毒H、F基因遗传多样性及其遗传进化分析

【目的】探究怀化地区犬瘟热病毒的基因变异与遗传多样性情况,阐明其遗传进化关系。【方法】从怀化地区收集30株犬源犬瘟热病毒样本,用PCR法扩增其H、F基因序列。利用DNAStar软件分析H、F基因序列碱基组成;利用Clustal X、MegAlign软件进行变异位点和遗传多样性分析;运用Clustal X、PhyML 3.0软件,采用最大似然法(ML)对怀化地区犬瘟热病毒进行遗传进化分析,并用FigTree v 1.3.1软件构建遗传进化树。【结果】30株怀化地区犬瘟热病毒均为Asia-Ⅰ型,其H、F基因序列长度分别为1778和1850 bp,其AT含量分别为56.8%~57.9%和54.9%~56.1%,GC含量分别为42.2%~43.0%和44.1%~44.9%。H基因的种内差异为0~3.1%,种间差异为51.36%~60.15%;F基因的种内差异为0~2.4%,种间差异为39.60%~53.09%。基于H、F基因序列所构建的遗传进化树发现,来源于怀化地区的30株犬源犬瘟热病毒全部位于遗传进化树的同一分支上。【结论】来源于怀化地区犬瘟热病毒之间虽存在一定程度的基因变异和遗传多样性,但它们之间的亲缘关系较近。且其H、F基因序列的种内差异小、种间差异大,是研究犬瘟热病毒基因变异、遗传多样性和遗传进化的重要遗传标记。

Cloning and Sequence of Glycoprotein H Gene of Duck Plague Virus

The glycoprotein H （gH） gene homologue of duck plague virus （DPV） was cloned by degenerate polymerase chain reaction （PCR） and sequenced. It was located immediately downstream from the thymidine kinase gene （TK）. In addition, the 3＇-end of the gene homologue to herpesvirus UL21 was located downstream from the gH gene. DPV gH gene open reading frame （ORF） was 2 505 bp in length and its primary translation product was a polypeptide of 834 amino acids long. It possessed several characteristics of membrane glycoproteins, including an N-terminal hydrophobic signal sequence, an external domain containing eight putative N-linked glycosylation sites, a C-terminal transmembrane domain, and a charged cytoplasmic tail. Comparison with other herpesvirus revealed identities of 20.2, 25.1, 23.0, 23.0, 26.5 and 26.0% with the gH counterparts of the human herpesvirus virus 1 （HSV1）, equine herpesvirus 4 （EHV4）, bovine herpesvirus 1 （BHV1）, pseudorabies virus （PRV）, gallid herpesvirus 2 （GHV2） and gallid herpesvirus 3 （GHV3）, respectively.

Research of AtNHX1 Gene Transformation in Brassica napus L. by Agrobacterium tumefaciens

[Objective] The aim was to investigate AtNHX1 gene transformation in Brassica napus L. mediated by Agrobacterium tumefaciens. [Method] By using Agrobacterium-mediated method and cre/lox plant expression vector,the transformation of AtNHX1 gene of Na＋/H＋ antiporter in Brassica napus was studied. [Result] The regeneration rate of cotyledon with petiole was much higher than that of hypocotyl,thus,the cotyledon with petiole was selected as the recipient for transformation. After the cotyledon with petiole was soaked in bacterial solution （OD600=0.4） for 8-10 min,kanamycin-resistant green seeding percentage could reach 3.75%. [Conclusion] The PCR detection of kanamycin-resistant plants proved that NHX1 gene had been inserted into Brassica napus genome. And this research could provide a new way to improve the salt tolerance of Brassica napus.