结构基因座和微卫星标记应用于绵(山)羊群体遗传分化的比较研究

对湖羊、同羊及长江三角洲白山羊的随机样本分别进行14个结构基因座和7个微卫星标记的遗传检测,比较由两种遗传标记获得的群体基因平均杂合度(H)、多态信息含量(PIC)及群体有效等位基因数(Ne);分别根据两种类型的基因频率资料,计算3个群体间的标准遗传距离并加以比较。结果表明:由微卫星等位基因频率获得的群体基因平均杂合度、多态信息含量及有效等位基因数显著大于结构基因座获得的,但在三群体中变化趋势是一致的;由结构基因座资料计算的3个群体间标准遗传距离为0 0268～0 2487,微卫星标记资料计算的3个群体间标准遗传距离为:0 2321～1 2313,绵山羊间显著大于绵羊群体之间。提示:结构基因座和微卫星标记一致揭示3群体内的遗传变异;同羊>湖羊>长江三角洲白山羊;微卫星标记较结构基因座标记更能表达近缘种间进化趋异水平,可将FCB11、MAF33、AE101、FCB128及FCB304位点作为研究绵山羊近缘种间遗传分化的标志性位点。     

全基因组水平蒺藜苜蓿反转录转座子IRAP分子标记开发及应用

由于苜蓿品种间遗传差异日益缩小,通过传统形态学鉴定表型愈加困难。在苜蓿育种过程中,利用分子标记可大大提高育种效率和品种鉴定。反转录转座子长末端重复序列（LTR）广泛分布在植物基因组中,基于LTR的分子标记具有丰富的多态性和高信息量等优势,被广泛用于物种品种鉴定、评价种质资源多样性等方面。本研究在全基因组水平鉴定和设计了大量蒺藜苜蓿LTR反转录转座子扩增多态性（IRAP）标记,并利用其对国内外40个紫花苜蓿种质资源进行遗传多样性分析。结果表明,根据设计开发出的431个IRAP引物,并按照其染色体位置信息组合获得69对IRAP引物。利用筛选出的37对多态性引物组合共扩增出325个等位位点,平均每个标记可产生8.8个等位位点;多态性条带比率（PPB）为50%~100%,平均值为79.9%;多态性信息含量（PIC）的范围为0.34~0.88,平均值为0.69。基于遗传相似系数（GS）对供试品种采用算术平均值非加权组平均法（UPGMA）进行聚类分析,以0.82为阈值可将40份供试材料分为4类,其分组结果与不同材料地理分布信息以及STRUCTURE分析相对一致。本研究首次在蒺藜苜蓿全基因组水平上开发了IRAP分子标记并在紫花苜蓿种质资源中进行评价与应用,获得的大量IRAP分子标记可对后续苜蓿品种的鉴定保护以及遗传背景分析提供技术支撑。     

广灵驴分子系谱的建立及群体遗传结构分析

旨在对广灵县优种驴场保种群体进行调查的基础上构建分子系谱,并对其种群的遗传结构进行分析。本研究采集保种群成年、体况良好的广灵驴（体重350~400 kg）颈静脉血10 mL（n=107）,其中公驴13份,母驴94份,抗凝处理后提取全血DNA。采用12个微卫星标记进行荧光PCR扩增后,用ABI3730测序仪进行分型。分型结果采用Cervus 2.0和Pedigraph 2.4软件构建分子系谱,同时采用STRUCTURE2.3和Fstat软件计算群体遗传参数,采用R语言的hclust函数绘制7头公驴及其后代的系统发生NJ树（邻接树）。结果,对107头种驴进行了分子系谱构建,找到了30头子代的父亲和7头子代的母亲,系谱可靠性>90%;微卫星标记的平均观测杂合度（H_Obs）和多态信息含量（PIC）分别为0.676 5和0.593 9,标记遗传多样性较高;NJ树对7个公驴家系进行了聚类;群体分化系数（F_ST）为0.184,群体平均近交系数（F_IT）为0.033,群体内近交系数（F_IS）为-0.238,且群体处于哈代-温伯格平衡状态,存在很弱的近交。本研究建立了广灵驴保种群可靠性较高的分子系谱并对其遗传结构进行了分析,证明该群体遗传多态性较高,群体近交系数较低,处于较好的保种状态,具有较大的品种资源开发潜力。     

2018-2020年中国部分省市猪流行性腹泻病毒S1基因的监测及遗传变异分析

【目的】 调查中国规模化猪场猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)的流行情况。【方法】 利用RT-PCR检测方法对2018-2020年采集于全国11个省市318个规模化猪场的2 391份样品进行PEDV核酸检测,对30份阳性样品进行S1全基因扩增与测序,并利用MegAlign、Mega 7.0等软件进行分析。【结果】 2018-2020年,猪场PEDV核酸阳性率分别为48.57%、10.96%和4.72%,样品PEDV核酸阳性率分别为28.97%、8.27%和7.50%,猪场和样品PEDV核酸阳性率呈现逐年下降的趋势。S1基因相似性分析显示,获得的30株毒株全部为GⅡ基因型,其中17株毒株为GⅡa基因亚型、5株毒株为GⅡb亚型、8株毒株为GⅡc亚型,GⅡa基因亚型毒株为2018-2020年流行的主要毒株类型;30株毒株S1基因序列之间核苷酸相似性为93.0%~99.5%, 氨基酸相似性为91.7%~100%,其中22株毒株与2017-2018年流行的GⅡ基因型毒株相比S1氨基酸序列表现出特征性插入和缺失。【结论】 本研究结果为监测和分析中国PEDV的变异和演化提供了临床数据,为猪流行性腹泻防控和疫苗研制提供了参考。     

牛至油博落回口服液的研制及体外抑菌活性测定

【目的】 研制牛至油博落回口服液, 并测定其体外抑菌活性及其主要成分的联合抑菌效果, 为临床用药提供理论依据。【方法】 通过预试验和Box-Behnken响应面法优化处方; 采用高效液相色谱法测定口服液主要成分含量; 采用滤纸片法测定口服液对大肠杆菌、沙门氏菌、金黄色葡萄球菌和粪链球菌的抑菌圈直径; 采用试管二倍稀释法测定口服液、5%牛至油溶液及1%博落回溶液对4种细菌的最小抑菌浓度(MIC)和最小杀菌浓度(MBC); 采用微量棋盘稀释法对5%牛至油溶液与1%博落回溶液进行体外联合药敏试验。【结果】 牛至油博落回口服液最优配方为: 5%牛至油, 1%博落回提取物, 25%增溶剂聚氧乙烯(40)氢化蓖麻油, 0.02%抗氧化剂2, 6-二叔丁基对甲酚(BHT), 余量为水。口服液中香芹酚的含量为42.59 mg/mL, 血根碱含量为6.51 mg/mL。口服液对4种菌的抑菌圈直径分别为16.9、16.4、23.7和17.0 mm, MIC分别为3.125、3.125、1.5625和1.5625 μL/mL, MBC分别为12.5、6.25、3.125和3.125 μL/mL; 5%牛至油溶液对4种菌的MIC分别为25、12.5、6.25和100 μL/mL, MBC分别为100、50、25和>200 μL/mL; 1%博落回溶液对4种菌的MIC分别为6.25、12.5、3.125和6.25 μL/mL, MBC分别为50、25、6.25和25 μL/mL。联合药敏试验表明, 二者联合用药对大肠杆菌、沙门氏菌起相加作用, 对金黄色葡萄球菌无作用, 对粪链球菌为协同作用。【结论】 试验制备了牛至油博落回口服溶液剂, 该制剂对大肠杆菌、沙门氏菌、金黄色葡萄球菌和粪链球菌具有良好抑制作用。     

Isolation,Identification, 16S rRNA Gene Sequencing and Genetic Evolution Analysis of Aeromonas aeruginosa from Duck

【Objective】 The experiment was aimed to study the pathogenic bacteria of acute death of ducklings in a duck farm in Henan province and its phylogenetic status.【Method】 The liver and spleen of dead ducks in the diseased duck farm were collected.The bacterial morphology, Gram-staining, biochemical characteristics, sequence analysis of 16S rRNA gene, drug sensitivity test and pathogenicity test were carried out.【Results】 The isolated bacteria grew on blood agar medium with smooth, convex, milky white and α-hemolytic Gram-positive cocci.Biochemical test results showed that the isolated bacteria were positive for maltose, sucrose, raffinose, nitrate reduction reaction, MP-VP test, urea, lysine decarboxylase, ornithine decarboxylase and aescin, and negative for xylose, lactose, glucose, sorbitol, hydrogen sulfide, citrate, peptone, mannitol, phenylalanine and methyl red.The results of 16S rRNA gene sequencing and BLAST comparison results showed that the 16S rRNA gene sequence of the isolated strain was 94.8%-99.9% similar to that of Aerococcus viridans published on NCBI, and the similarity with strains Mnlv1, Mnlv2 and W66 was the highest, up to 99.9%.The similarity with strain GXBl-1 was the lowest, which was 94.8%.Phylogenetic tree analysis showed that the isolated bacteria belonged to the same genus and group on the same branch as strains of Aerococcus viridans FL09, 15MS and Mnlv2 etc., and had the closest genetic relationship with FL09.The results of drug sensitivity showed that the isolated strain was sensitive to fosfomycin, ampicillin and amoxicillin, moderately sensitive to penicillin, neomycin and amikacin, it was resistant to rifampicin, neomycin, bacitracin, erythromycin and enrofloxacin.The results of pathogenicity test showed that the strain was lethal and increased with the increase of inoculation concentration.【Conclusion】 This study determined that the pathogen causing the acute death of ducklings in a duck farm in Henan province was Aerococcus viridans, which provided a reference basis for clinical diagnosis and treatment of the diseases caused by this bacteria.     

猪多杀性巴氏杆菌的分离鉴定及oppA基因遗传进化分析

【目的】了解福建省猪场猪多杀性巴氏杆菌（Pasteurella multocida,Pm）的流行及oppA基因遗传进化情况。【方法】本研究采用细菌分离培养、生化试验、16S rRNA PCR扩增测序、PCR荚膜分型、oppA基因克隆及相似性分析、动物回归试验等方法对分离菌株进行鉴定和分析。【结果】本研究共分离到10株菌,分离菌在血平板上形成淡灰白色、湿润光滑、奶油露珠状菌落;分离菌株能酵解蔗糖、果糖、麦芽糖和甘露醇,不能分解葡萄糖、枸橼酸盐、乳糖、硫化氢等,与多杀性巴氏杆菌生化特性基本一致;分离菌株16S rRNA序列与GenBank中登录的多杀性巴氏杆菌相似性达99.9%以上,10株分离菌均为多杀性巴氏杆菌;PCR荚膜分型显示,6株分离菌为荚膜A型,4株为荚膜D型;基于oppA基因的遗传进化树显示,10株分离菌均位于同一分支内;动物回归试验结果显示,在24 h内攻毒小鼠死亡率较高（21/30）,分离菌有较强的致病力。【结论】福建省猪场猪多杀性巴氏杆菌流行菌株的荚膜血清型主要是A和D型,且大部分菌株都来源于共同的祖先,本研究结果丰富了猪多杀性巴氏杆菌的流行病学资料,并为该病的防控奠定基础。     

Investigation of Co-infection of PPV7 and PCV2 and Genetic Variation Analysis for PPV7 Cap Gene in Fujian and Guangdong

【Objective】 To investigate the co-infection situation of Porcine parvovirus (PPV7) and Porcine circovirus type 2 (PCV2)in Fujian and Guangdong, and to understand the molecular genetic characteristics of PPV7 Cap gene.【Method】 The blood sample of 432 infected pigs from 69 pig farms in Fujian and Guangdong were collected to detect PPV7 and PCV2 by PCR.The PPV7 Cap gene of positive samples was cloned and sequenced.The DNAStar software was used to analyze the nucleotide and amino acid sequences of PPV7 Cap gene, and Mega 7.0 software was used to draw the genetic evolution tree.【Result】 The results showed that the positive rate of PPV7 was 21.99% (96/432), the positive rate of farms was 53.62%(37/69), and the positive rate of PCV2 was 54.17% (234/432), and the co-infection rate of PCV2 and PPV7 was 13.43% (58/432).The 17 PPV7 Cap gene sequences were amplified using PCR.Nucleotide homology analysis revealed that the homology of the 17 PPV7 Cap gene sequences was 85.6%-100%, and the homology with reference strains was 85.8%-99.0%.Amino acid sequence comparison analysis revealed that the amino acid homology of the 17 PPV7 Cap protein sequences was 87.6%-100%, and the amino acid homology with reference strains was 82.6%-98.7%.Phylogenetic analysis of Cap gene showed that PPV7 could be divided into five main evolutionary branches of PPV7a-PPV7e, among which 9 isolates belonged to PPV7a subtype, 3 isolates belonged to PPV7b subtype, 4 isolates belong to PPV7c subtype, and only 1 isolates isolates belonged to PPV7e subtype.【Conclusion】 This study indicated that PPV7 was widely prevalent in Fujian and Guangdong regions, and had a high co-infection rate with PCV2, which might be the pathogenic factor of Porcine circovirus associated disease(PCVAD).The genetic diversity of PPV7 isolates was abudant in both regions, and PPV7a was the dominant strain at present.The findings of this study provided theoretical basis and data reference for PPV7 prevention and control and vaccine research.     

Research Advancement in Genetic Variation of Wood Properties

The genetic improvement of wood properties is one of the important measures of improving wood quality. To achieve that, forestry and wood scientists have done a great deal of research in expectation to produce more high-quality wood. The paper summarizes the researches on genetic variation and genetic correlation of wood properties and proposes the prospects to conduct the genetic improvement of wood properties.     

滇杨遗传改良策略初论

滇杨是我国西南地区特有的一种乡土树种，也是我国乃至世界少有的分布于低纬度高海拔地区的杨属树种。针对目前国内滇杨遗传改良研究几为空白的现状，对其遗传改良的途径进行了系统探讨。从滇杨的资源特点及研究现状出发，在借鉴国内外杨树遗传改良成功经验的基础上，提出常规育种与现代生物技术育种有机结合，形成一个系统高效、合理可行的滇杨遗传改良策略。