溶藻弧菌单克隆抗体的制备及应用

小鼠骨髓瘤细胞系SP2/0与经溶藻弧菌(1.1587)免疫的BALB3/C雌鼠脾细胞在PEG条件下融合,有45.8%的培养孔有杂交瘤细胞生长,其培养上清液抗体阳性率为77.4%.经反复有限稀释法克隆杂交瘤细胞,获得7株抗溶藻弧菌的单克隆抗体杂交瘤细胞株,分别为AC1-C9,AC1-C11,BB4-D4,AD1-A3,AD1-F3,AD2-E7,AD2-F7.取AD1-F3扩大培养,注射小鼠,制备了抗溶藻弧菌的单克隆抗体腹水,滴度为11000.该腹水与其他3株溶藻弧菌菌株有强交叉反应,与其他13株标准菌株无交叉反应.利用制备的单抗腹水,建立了检测溶藻弧菌的单抗-ELISA技术.该反应系统可应用于溶藻弧菌的快速诊断,反应时间为6-7h,检测灵敏度为104cells·-1.并利用该检测方法进行了11份待测菌样本溶藻弧菌的检测.     

用PCR法快速筛选中国对虾含微卫星的重组阳性克隆

运用PCR在中国对虾的小片段基因组文库中快速筛选含有微卫星序列的重组阳性克隆。PCR中使用的引物为载体pGEM-3(zf( )上多克隆位点两侧的T7／Sp6启动子引物和根据微卫星核心重复序列设计的STR引物：STR1：5‘-ATATATATATAT－3‘：STR2：5’－TCTCTCTCTCTC－3’，直接使用含有重组克隆的菌液，在PCR反应前增加一段裂解细菌的高温。筛选出来的含有微卫星序列的重组阳性克隆经DNA测序验证。结果证明，采用PCR方法用菌液快速筛选含有微卫星序列的重组阳性克隆完全可行，而且具有简便、快速、高效、不涉及同位素操作等优点。     

传染性脾肾坏死病毒_ConA和L_省略_对体外鳜的头肾淋巴细胞转化的影响

采用MTT颜色反应法研究传染性脾肾坏死病毒（ISKNV）、伴刀豆球蛋白A（concanavallin A,ConA)和脂多糖(lipopolysaccharide,LPS）在离体状态下对健康和ISKNV感染后存活30d或60d的鳜的头肾淋巴细胞转化的影响。结果表明，ConA和LPS能促进健康鳜头肾淋巴细胞的转化，而对感染后存活30d或60d的鳜头肾淋巴细胞没有作用；纯化的ISKNV对健康鳜头肾淋巴细胞没有促进淋巴细胞转化的作用，但对ConA的作用有抑制，对感染后存活的30d或60d的鳜头肾淋巴细胞，纯化的ISKNV有一定的促进转化的作用，感染存活的鳜头肾中有对ISKNV的免疫记忆性T细胞。另外，用建立的ISKNV PCR检测方法对健康和ISKNV感染后存活的鲜组织进行了检测，结果显示，健康鳜为阴性，而ISKNV感染后存活的鱼，头肾、后肾、脾脏和部分鱼的心脏为阳性。ISKNV在鳜体内形成潜伏感染。     

真鲷消化道的组织学和形态学研究

采用扫描电镜和光镜技术对真鲷的消化道进行了组织学和形态学研究。具发达颌齿的口咽腔之后，真鲷消化道由分化明显的食道、胃、小肠和直肠构成。口咽腔粘膜表面为复层上皮，其中含有怀状细胞和味蕾，表层细胞具微褶（脊）；食道粘膜也衬着复层上皮，由表层具微褶（脊）的扁平细胞、中层的怀状细胞及底部的基底细胞组成，食道粘部分区域 衬着单层柱状上皮；胃Ｖ形，粘膜表层为单层柱状上皮，贲门部和盲囊部上皮下有一厚层胃腺组织；     

从中国对虾ESTs中筛选微卫星标记的研究

利用生物信息学方法在含有10446个中国对虾ESTs的数据库中进行微卫星序列的筛选，共发现微卫星序列229个，占整个ESTs数据库的2．19％，其中含双碱基重复序列146个和3碱基重复序列58个，分别占在ESTs数据库中发现微卫星序列总数的63．76％，和25．33％，大部分发现的微卫星序列均为Perfect形式的重复序列。根据筛选得到的微卫星序列设计并合成引物19对进行多态性检测，在有扩增产物的16对引物中，首次筛选得到8个中国对虾微卫星标记，并对这些微卫星标记进行了等位基因频率、观测杂合度、期望杂合度、PIC值等统计学指标的评价。结果表明，在8个微卫星位点上，等位基因的数目从5到15不等，等位基因长度从：165～305bp，期望杂合度和多态性信息含量分别为0．59到0．89和0．56到0．88，表明这8个中国对虾微卫星标记完全适合于遗传分析。     

应用栖息地指数对印度洋大眼金枪鱼分布模式研究

运用分位数回归方法对温度、温差、氧差与印度洋大眼金枪鱼延绳钓钓获率进行二次回归分析,找出最佳上界方程,以最佳上界方程拟合的数值来建立栖息地指数(HSI)模型,从而揭示印度洋大眼金枪鱼栖息地的分布模式。研究表明,温度、温差、氧差与印度洋大眼金枪鱼延绳钓钓获率的最佳上界分位数回归方程分别为HR_(T0.9)=-44.803 7.685T_(0.9)-0.255T_(0.9)~2,HR_(dT_(0.9))=6.234 0.953dT_(0.9)-0.026dT_(0.9)~2和HR_(dO_(0.88))=7.422 4.25dO_(0.88)-0.727dO_(0.88)~2。10°N～10°S间印度洋海域大眼金抢鱼HSI指数达到0.9以上;10°N以北的波斯湾及10°S～15°S海域的HSI指数为0.8～0.9;15°S～40°S之间海域HSI指数介于0.7～0.8,其中50°E～90°E、15°S～25°S间存在一片季节性HSI指数<0.7的区域;40°S以南的海域HSI指数<0.6。     

拟穴青蟹GRP78基因的克隆与应激表达

葡萄糖调节蛋白78(glucose regulated protein 78 ku,GRP78)是热休克蛋白70家族成员之一,在调节蛋白质折叠和维持内质网稳态过程中起着分子伴 侣作用。采用RT-PCR、RACE等技术,首次从拟穴青蟹获得了GRP78的cDNA全长序列。该序列全长2 284 bp,开放阅读框(ORF)为1 962 bp,编码653个氨基酸残基。 同源分析显示,该基因编码的蛋白含有HSP70家族的签名序列,C末端为内质网蛋白滞留信号KDEL,与其他物种具有很高相似性。实时荧光定量PCR结果表明,GRP78 基因在拟穴青蟹多个组织中均有表达。第一期仔蟹在不同的温度和盐度下暴露12 h后,GRP78基因表达量随环境温度升高而增加;在高盐(30)条件下GRP78表达量 较高,进而推测拟穴青蟹GRP78参与蛋白质折叠和环境胁迫的应答。     

Einfluss selektiver mechanischer Fruchtbehangsregulierung auf Ethylensynthese als Stressindikator sowie Ertrag und Fruchtqualität bei Kernobst

The overall objective of this work was to improve fruit quality, break alternate bearing and reduce hand thinning using fewer chemicals in fruit crops. A device was constructed for mechanical thinning, which consisted of three independent horizontal rotors with ropes and freely adjustable angles on a frame, mounted on a front three point hitch and powered by the tractor hydraulics. This can be adapted to any fruit tree trained as spindle, Solaxe, (tall) vertical axis or fruit wall (le mur fruitier) irrespective of rootstock employed. Rotor speed varied from 300 to 460?rpm at either 5 or 7.5?km/h tractor speed. Eight-year-old or twelve-old apple trees cvs. ‘Gala’ and ‘Golden Delicious’ were mechanically thinned in 2007 between pink bud and full bloom (flower bud stages 6–8 or F1–F2) near Bonn, Germany; non-thinned and hand-thinned apple trees of the same block and variety served as control. Mechanically thinned flowering branches showed a similar amount of ethylene efflux (0.4–0.6?ppm C₂H₄/branch) as non-thinned flower branches, preventing potentially unexpected subsequent fruit drop, except for those removed by the rotors. The impact of the horizontal rotors on the branches was from the upper side and removed excessive flowers right to the tree trunk viz. the centre of the tree canopy, where fruits of lesser quality are expected leaving 2–3 flowers per cluster. Leaf damage was less than??10%, even at the fast rotor speed of 420?rpm, which was associated with negligible wood injury. Mechanical thinning induced firmer and sweeter fruit, i.e. tastier apples with longer shelf life, relative to control fruit from non-thinned apple trees. The greatest efficacy in terms of final fruit quality in the grading/sorting was achieved by a rotor speed of 360?rpm at a tractor speed of 5?km/h: Fruit mass increased by up to 20?g and the proportion of fruit larger than 70–75?mm by 10–30% compared with the fruit from non-thinned trees. Mechanical thinning with this newly constructed device led to a 10–20% reduction in yield, but increased returns due to better fruit size and colouration in apple with the potential to overcome alternate bearing.     

Molecular identification and genetic analysis for 24 turf-type Cynodon cultivars by Sequence-Related Amplified Polymorphism markers

Bermudagrass (Cynodon ssp.) germplasm is genetically diverse and widely distributed in the world. The study was conducted to identify and assess the molecular variation and relationship among 24 cultivars developed in China, Australia and the USA. Sequence-Related Amplified Polymorphism (SRAP) was applied to cultivars identification in this study for the first time. Thirty of the 90 SRAP primer combinations generated a total of 274 clearly bands encompassing 249 (91%) polymorphic. Each bermudagrass cultivar has its unique binary code and can be distinguished from the others. Three distinct clusters were obtained by unweighted pair-group method with arithmetic averages based on the polymorphic markers. Coefficients of genetic distance among the genotypes ranged from 0.57 to 0.97. The results demonstrated that SRAP marker is a stable molecular marker technique for the identification of bermudagrass cultivars and their genetic relationships.     

白菜亚硝酸还原酶基因BcNiR的克隆及表达分析

 以白菜品种‘苏州青’自交系叶片cDNA为模板, 采用RT-PCR、3′RACE和5′RACE技术, 获得了编码亚硝酸还原酶基因(NiR) 的cDNA全序列1 852 bp, 包含有1 749 bp的开放阅读框, 编码583个氨基酸, 命名为BcNiR。所推导的氨基酸序列与拟南芥NiR1、烟草nii2编码的氨基酸序列具有较高同源性, 分别为83%和76%。生物信息学分析表明, BcNiR具有完整的NiR蛋白结构, 含血红素蛋白β - 化合物区域, 一个明显的西罗血红素siroheme结合位点和4Fe-4S区域, 可以在SWiSS-MODEL数据库中搜索到与之相近的三维结构。RT-PCR结果显示, 该基因在叶片中的表达量远远高于根系, 在以0、10、20、30、40 mmol·L ^{- 1}硝态氮各分别处理0、2、4、6、8、12 h的试验中, 30 mmol·L ^-1处理4 h可使BcNiR的表达量达到最大; 5和10 mmol·L ^{- 1}铵态氮处理试验表明, 高浓度的铵抑制该基因的表达。