一株鸭源禽流感病毒（H9N2）全基因序列分析及其排毒规律的研究

为研究一株鸭源H9N2亚型禽流感病毒(JN1株)全基因组的分子特征和遗传进化情况以及感染雏鸭后的排毒规律,对其生物学特性、全基因组序列以及遗传进化进行了分析,通过点眼滴鼻方式感染3周龄健康雏鸭,利用建立的实时荧光定量PCR(RT-PCR)检测鸭的排毒规律,并测定血清抗体效价。JNI株的鸡胚半数感染量为10~(-7.54)/0.2mL,鸡胚最小致死量的平均死亡时间为72h,静脉致病指数为0.266,抗原相关系数为0.47。系统发育分析表明,JN1株的HA与CK/HK/G9/97和DK/HK/Y280/97在同一分支上,NA、M、NP、PA、PB1、PB2基因与SH/F/98同源性较高,NS基因则与H5N1亚型禽流感病毒进化关系较近。HA的裂解位点为PSRSSR↓GL,存在6个潜在糖基化位点,在234位的氨基酸残基为L,NA基因颈部存在缺失。雏鸭在感染该病毒后,饮食正常,精神良好,3~17d均有排毒,咽拭子和泄殖腔棉拭子排毒规律基本一致,分别在第3天和第13天出现排毒高峰。该H9N2亚型禽流感病毒属欧亚大陆分支,为低致病性禽流感,病毒可通过呼吸道和泄殖腔向外界排毒,排毒时间较长。     

小黑麦与黑麦花序结构和籽粒特性比较

植物的花序结构及其形态特征是影响种子生产性能的重要因素。对于禾本科植物而言,其花序大小、小穗数、小花数及小花的外稃、内稃和芒的性状特征直接影响种子形成和产量。本研究通过测定小黑麦品系C2、C35和黑麦品系C13、C33的花序结构特征,得到以下结果:小黑麦花序[(14.30±0.52) cm×(1.24±0.09) cm]明显大于黑麦[(13.20±0.35) cm×(0.82±0.02) cm],小黑麦花序长而粗,黑麦花序短而细;小黑麦花序的小穗数[(21.35±1.47)个]少于黑麦[(30.20±0.79)个];小黑麦每个小穗的小花数较多,为3~4朵,黑麦的小花数趋于稳定,每个小穗有2朵小花。从花序中部小穗的小花结构看,小黑麦下位护颖长[(1.27±0.11) cm]和宽[(0.26±0.03) cm]、上位护颖长[(1.30±0.09) cm]和宽[(0.23±0.04) cm]均显著大于黑麦下位护颖长[(1.04±0.05) cm]和宽[(0.08±0.01) cm]、上位护颖长[(0.94±0.10) cm]和宽[(0.06±0.01) cm];小黑麦中部小穗第1小花的外稃宽[(0.32±0.03) cm]、外稃高[(0.24±0.03) cm]和芒长[(8.35±0.51) cm]、第2小花的外稃宽[(0.35±0.04) cm]、外稃高[(0.25±0.05) cm]和芒长[(8.37±1.19) cm]极显著大于黑麦相应值[(0.26±0.01),(0.15±0.01),(5.50±0.19),(0.25±0.01),(0.17±0.01)和(5.18±0.23) cm];第1和第2小花的外稃长[(1.35±0.06),(1.37±0.06) cm]和内稃长[(0.84±0.04),(1.41±0.06) cm]均小于黑麦的外稃长[(1.49±0.05),(1.47±0.05) cm]和内稃长[(1.45±0.05),(1.47±0.04) cm]。小黑麦的穗粒数[(52.50±1.80)粒]显著低于黑麦[(58.50±2.50)粒] (P<0.05),但其穗粒重[(2.08±0.04) g]、粒重[(0.04±0.00) g]和籽粒宽[(3.04±0.32) mm]均极显著高于黑麦 (P<0.01),每个花序的籽粒不仅体积大,而且质量较重,其籽粒的生产性能高于黑麦。小黑麦籽粒呈椭圆形或长卵圆形,浅黄色,表皮皱缩,饱满度差;黑麦籽粒呈窄纺锤形,青灰色,表皮光滑,籽粒饱满。小黑麦和黑麦花序结构与籽粒性状的Pearson相关分析表明,小黑麦花序宽和花序基部小穗数与籽粒长显著正相关(P<0.05),花序长与籽粒宽极显著正相关(P<0.01),花序中部小穗的下、上位护颖宽分别与穗粒重和穗粒数极显著正相关(P<0.01);黑麦花序中部小穗第2小花的内稃长与籽粒长显著正相关(P<0.05),外稃高与穗粒数极显著负相关(P<0.01)。研究小黑麦和黑麦的花序结构和籽粒特征,对正确区分二者和了解其籽粒的生产性能具有重要意义,同时有利于小黑麦的示范推广。     

草地群落类型对乳白香青种群特征和雌雄株形态及抗性的影响

为探究亚高山雌雄异株草本植物乳白香青对生境的适应方式,通过测定5个草地群落(A-老芒麦改良草地,B-虉草改良草地,C-天然草地,D-天然草甸,E-沙化草地)中乳白香青种群数、种群密度和面积、分布系数、性比组成,以及雌雄株构件性状、生物量配比、水分分配及抗性物质含量等参数,研究群落组成对乳白香青种群特征、雌雄株形态可塑性及抗性的影响。结果表明,5个草地群落的优势种、群落密度、基盖度、物种密度和优势种高度及土壤理化性状的显著差异(P<0.05),使不同群落中乳白香青种群特征显著不同,群落类型对种群特征参数影响顺序为种群面积>种群密度>性比>年龄结构>种群数>分布系数;D群落中乳白香青种群数和分布系数最高,C中乳白香青种群面积和密度最大,E中乳白香青种群数仅分别为D和C中的30.21%和35.11%,分布系数仅为C中的36.04%;群落类型对乳白香青种群年龄结构和性比有显著影响(P<0.05),雌株数为雄株的2.22~7.03倍;叶数和叶面积、株高和茎径、根长、小花数和苞片层数有显著的性别差异(P<0.05),群落类型对雌雄株的叶数和叶面积、株高、根数、花序数和小花数有显著影响,对雄株茎径和苞片层数有显著影响;雄株总生物量和花分配比显著高于雌株,而根分配比显著低于雌株(P<0.05),群落类型对乳白香青生物量配比有显著影响,雌株生物量配比受影响顺序为花>总生物量>叶>根>茎,雄株受影响顺序为花>茎>叶>总生物量>根;雄株茎、叶、花含水量显著高于雌株(P<0.05),根含水量在性别间和群落间差异较小,雌株构件含水量受群落类型影响顺序为花>茎>叶>根,雄株为叶>茎>花>根。雄株中MDA含量高于雌株,雌株中H₂O₂含量及T-AOC活性大于雄株(P<0.05),群落组成对雌、雄株中H₂O₂和MDA含量及T-AOC活性有极显著影响(P<0.01)。A和B中H₂O₂和MDA含量最低,E中H₂O₂和MDA达最大值,而T-AOC活性最低。雌、雄个体均能依据群落生境调整生长策略,通过构件性状、生物量配比、水分分配和抗性生理等适应性变化,使种群具有与生境匹配的种群特征,利于提高种群适合度和生活史完成。     

羊草种子萌发相关基因的筛选及表达分析

前期实验表明,变温处理(28 ℃ 12 h/16 ℃ 12 h)可显著提高羊草种子的萌发率,且羊草种子萌发中的第1天是接受变温信号的关键时期。以此为研究基础,结合羊草种子变温萌发的转录组测序数据,针对羊草种子萌发初期对变温处理的响应筛选出与种子萌发、休眠及低温相关的基因24个,利用测序结果中这些基因的RPKM值制作基因表达热图并分析其表达差异。以萌发率高、低的两种羊草种质的种子为材料,对24个基因在恒温12 h(28 ℃)和变温1 d(28 ℃ 12 h/16 ℃ 12 h)萌发处理中的表达分别进行了定量分析。结果表明,与恒温对照相比,变温处理12 h后,表达明显上调的基因有SAIN1,PP2C62,EXPB3,EXPB4,GA3ox,EXPA2和EXPA7,而表达明显下调的基因有bHLH49,GID1,ABI8,Chi1,11833,CBF3,NAC2,PP2C72,SAIN2和5423。通过进一步分析相关基因在高、低萌发率两个种质中表达的差异,筛选出其中可能与羊草种子萌发相关的基因有几丁质酶基因Chi1,转录因子基因CBF3,羊草新基因5423,赤霉素合成基因GA3ox,细胞松弛素蛋白基因EXPB4和羊草新基因SAIN1,将为下一步阐明羊草种子萌发的分子作用机理奠定基础。     

低温胁迫对狗牙根生理及基因表达的影响

以耐寒性差异较大的杂交狗牙根品种运动百慕大、天堂419和普通狗牙根品种保定狗牙根为试验材料,分析了昼夜温度为适温(30 ℃/25 ℃)、亚适温(18 ℃/12 ℃)、冷害(8 ℃/4 ℃)和冻害(4 ℃/-4 ℃)4种温度处理下,低温对狗牙根生长速率和草坪质量、MDA含量、叶绿素荧光(F_v/F_m)、光合作用(P_n、G_s、C_i、T_r)、H₂O₂和O2·-含量、抗氧化酶活性(SOD、POD、CAT、APX)、抗氧化酶及耐寒相关基因(CBF1、COR、LEA)表达的影响。结果表明,随着温度的降低,狗牙根草坪质量、生长速率、F_v/F_m和光合作用显著下降,但在耐寒性强的运动百慕大中下降幅度较小;低温下叶片MDA和H₂O₂含量及O2·-产生速率显著升高,其中在耐寒性弱的保定狗牙根中升高程度更大;狗牙根叶片抗氧化酶活性及抗氧化酶基因的表达水平随着处理温度的下降而显著升高,其中在运动百慕大中更为明显;狗牙根CBF1、COR和LEA基因表达量随着温度的下降而显著升高,尤其是在耐寒狗牙根运动百慕大中更为显著。低温诱导的抗氧化酶和耐寒相关基因上调表达,增强了细胞内的抗氧化防御,有助于狗牙根植株维持较高的光合作用和细胞膜稳定性,延缓叶片的衰老,从而提高了狗牙根的抗寒能力。     

Differential gene expression of Eph‐ephrin A1 and LEPR‐LEP with high or low number of embryos in pigs during implantation

The objective of this study was to ascertain whether mRNA and protein expressions of implantation‐related genes (erythropoietin‐producing hepatocellular receptor–ligand A1, Eph‐ephrin A1 and leptin receptor–leptin, LEPR‐LEP) differed between pigs with high and low number of embryos, and whether these differences in gene expression might affect embryo implantation. Experimental pig groups (n = 24) for high and low number of embryos were prepared by altering the number of eggs ovulated in pre‐pubertal gilts treated with 1.5 × (High) or 1.0 × (Low) PG600 ([400 IU PMSG + 200 IU hCG]/dose, AKZO‐NOBEL). Gilts expressing oestrus were artificially inseminated twice and maintained in breeding and gestation until the reproductive tract was collected on day 22 of pregnancy. At slaughter, the reproductive tracts from each pregnant gilt from each treatment were immediately processed to collect samples for RNA and protein analysis. Within each gilt, three conceptus points were sampled, one from each horn and then a random conceptus within the tract. At each conceptus point, endometrial attachment site, chorion–allantois and embryo were collected and immediately frozen in liquid nitrogen. Number of corpus luteum (CL) (35.4 vs. 12.6) and total embryo number (18.8 vs. 10.2) were greater in the high‐embryo compared to the low‐embryo group, respectively (p < .05). Real‐time qPCR results showed that Eph‐ephrin A1 mRNA expression was less in the high‐embryo (p < .05) compared to the low‐embryo group. In addition, Western blotting analysis indicated that Eph‐ephrin A1 and LEP protein expression at endometrial attachment site in high‐embryo was less (p < .05) compared to low‐embryo group. It was also noted that mRNA expression of Eph‐ephrin A1 and LEPR‐LEP was greater in pregnant than non‐pregnant gilts (p < .05). Moreover, mRNA expression of Eph‐ephrin A1 (p < .05) and LEPR‐LEP was greatest at endometrial attachment site among all three tissues. There was a positive correlation between expressions of Eph‐ephrin A1, LEPR‐LEP and embryo length with the correlation coefficient 0.31–0.59. For Eph‐ephrin A1, the highest correlation coefficient appeared between Eph A1 expression and normal embryo number, between ephrin A1 expression and embryo length. For LEPR‐LEP, the highest correlation coefficient appeared between LEPR‐LEP expression and ovary weight (0.79 for both, p < .05), followed by embryo length and weight. The results of this study suggest that low expression of Eph‐ephrin A1 and LEPR‐LEP is somehow related to increased embryo number during implantation and that endometrial attachment site might be the main target tissue of these gene products. Yet, the increased expression of Eph‐ephrin A1 and LEPR‐LEP appeared associated with increased embryo growth (length and weight) and ovary weight, Eph‐ephrin A1 and LEPR‐LEP might play roles in the regulation of embryo implantation in pigs.     

Effects of five cryoprotectants on proliferation and differentiation‐related gene expression of frozen‐thawed bovine calf testicular tissue

The cryopreservation of testicular tissue is a potential method for preserving male fertility. However, the effect of cryopreservation on bovine calf testicular tissue is scarce. This study investigated the effect of different cryoprotectants on bovine calf testicular tissue at the molecular level. Testicular tissue from ten immature bovine calves (6 months) was collected after slaughter and cryopreserved in an extender containing different concentrations of the following five cryopreservation solutions (CP): bovine serum albumin (BSA) with 5% dimethyl sulfoxide (DMSO), trehalose with 5% DMSO, DMSO and glycerol and ethylene glycol (EG). After 7‐day cryopreservation, the expression levels of three spermatogonial stem cell (SSC)‐related genes, octamer‐4 (OCT4), KIT ligand (MGF/SCF) and kit oncogene (C‐KIT), were investigated by quantitative PCR (qPCR). The cell viability was highest for the tissues preserved with 30 mg/ml BSA (77.82% ± 1.22) and 40 mg/ml trehalose (74.23% ± 1.16) compared with other groups (p < 0.05), and the level of expression of the three genes was highest with 30 mg/ml BSA (p < 0.05). Compared with other CPs, the 30 mg/ml BSA and 40 mg/ml trehalose have the better cryopreserve protection. The 30 mg/ml BSA is the most viable media for the cryopreservation of testicular tissue from cattle.     

基于冗余分析的高寒草原土壤与草地退化关系

在高寒草原退化草地设置研究样地,观测植物群落特征、采集土壤样品、分析土壤物理化学性质,依据数量生态学基本原理,探讨高寒草原退化草地与土壤因子间关系。结果表明:不同退化程度样地沿冗余分析排序图第一排序轴分布,第一排序轴反映草地退化程度的变化;草地植物群落中与第一排序轴负相关且按相关程度大小排序的指标为盖度>地上生物量>地下生物量;第一、第二排序轴能够解释97.1%的土壤因子与草地退化关系;土壤沙砾、p H与第一排序轴正相关,土壤有机碳、土壤含水量、容重、全氮、有效氮与第一排序轴负相关;第一排序轴及所有排序轴所反映的土壤因子均与草地退化之间呈极显著相关关系(P<0.01);不同土壤因子与草地退化之间相关程度不同,土壤有机碳(r=-0.890)、土壤含水量(r=-0.864)、容重(r=-0.847)、全氮(r=-0.836)、有效氮(r=-0.821)等与高寒草原退化相关程度更高且相关关系极显著(P<0.01)。利用退化草地群落特征和土壤因子数据矩阵进行冗余分析能够较好地综合反映土壤因子与草地退化之间的关系及相关程度,土壤有机碳、土壤含水量、容重、全氮、有效氮是反映高寒草原退化的重要土壤指标。     

叶面喷施吲哚乙酸对油菜蕾薹期渍水的缓解效应

以油菜品种秦优7号(敏感)和苏油4号(耐渍)为材料,通过盆栽试验研究吲哚乙酸(IAA)对油菜蕾薹期渍水的缓解效应。蕾薹期渍水持续10d,渍水结束后,叶面喷施浓度为0.1mmol/L的IAA,随后的0~15d调查油菜的生理变化。结果发现:渍水后生物量和叶片光合色素含量降低;谷胱甘肽、抗坏血酸含量和超氧化物歧化酶活性升高;丙二醛含量升高;蔗糖合成酶、蔗糖磷酸合成酶活性上升,蔗糖和可溶性糖含量增加。渍水结束后喷施IAA,对敏感的秦优7号缓解效应更明显;叶片中的光合色素含量升高,5d后显著恢复生长;蔗糖磷酸合成酶活性降低,降低蔗糖含量;蔗糖合成酶活性降低,减少蔗糖的分解;尤其是喷施15d后IAA对渍水胁迫的缓解作用最明显,苏油4号根干重显著增加,而秦优7号的根干重和茎干重均显著增加。叶面喷施IAA有效缓解单株角果数、每角粒数和产量的降低,但对抗氧化系统影响较小。     

一种准零刚度被动隔振系统的非线性共振响应分析

分析了一种机构的负刚度特性和它与线性主弹簧并联后而成的隔振系统的准零刚度特性，推导了准零刚度被动隔振系统的主谐波共振、亚谐波共振和超谐波共振的幅频特性表达式.结果表明，机构的负刚度区域随弹簧的初始变形与杆长的比值增大而变宽；隔振系统的准零刚度区域随主弹簧的刚度与机构中的弹簧总刚度的比值以及机构中的弹簧初始变形与杆长的比值增大而变宽；经比较发现，本文推得的被动隔振系统的同一种谐波共振幅频特性与杜芬方程的幅频特性均存在较大偏差.因此，对于这类被动隔振系统的运动微分方程，若以位移激励的幅值与激励频率平方的乘积代替单位质量的激励力幅值按杜芬方程的解来处理谐波共振问题将会带来较大误差.