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。     

太平洋牡蛎(Crassostrea gigas)类α-1,2-岩藻糖基转移酶的密码子优化与原核表达

牡蛎消化组织内存在的类A型血型组织抗原是其特异性富集诺如病毒的主要原因,FUT2(Fucosyltransferase 2,α-1,2-岩藻糖基转移酶)是A型血型组织抗原合成的关键酶.本研究在前期克隆了太平洋牡蛎(Crassostrea gigas)类FUT2基因cDNA全长的基础上,根据大肠杆菌密码子偏爱性优化并合成了类FUT2基因,插入原核表达载体pRSET A构建pRSET-mof,将其转化大肠杆菌BL21,用异丙基-β-D-硫代半乳糖苷(IPTG)诱导.经SDS-PAGE分析显示,在37℃、IPTG终浓度为0.8 mmol/L的条件下,诱导4h后出现大小约为46 kDa的特异性目的条带.利用His亲和层析柱纯化及超滤管浓缩目的蛋白,得到单一条带,说明纯化效果良好.Western blot分析显示,目的蛋白与抗6×His标签单克隆抗体、抗人FUT2单克隆抗体均能发生特异性反应,表明优化后的太平洋牡蛎类FUT2基因在大肠杆菌系统中成功表达.本研究结果为今后研究太平洋牡蛎类FUT2基因的功能,进一步探索牡蛎特异性富集诺如病毒的分子机理奠定了基础.     

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.     

Processing tomato quality as affected by irrigation scheduling

In Southern Italy the paucity of water is a very important problem and due to the fundamental importance of the water for the processing tomato, in this study the influence of irrigation on processing tomato quality was investigated in the years 2002 and 2003. A hybrid processing tomato (Lycopersicon esculentum L. Mill cv. Ability) was treated with different water regimes obtained combining amount of water and irrigation interval. The effects of the trials were evaluated taking into account the physical and chemical characteristics of the fruits, as well as the content in antioxidants moieties. Furthermore, the relationship among all the quality parameters and the seasonal amount of irrigation water was evaluated.The results, comparing different water regimes obtained combining amount of water and irrigation interval, gave useful indications on the possibility to improve nutritional tomato quality by reducing irrigation water during tomato cropping. In particular, relatively to the Mediterranean areas of tomato cultivation and to the texture of the soil considered in this study, the best compromise between quality and quantity of the processing tomato fruit was achieved both with the extension of the irrigation interval (L40 or L60) and with the limitation of irrigation volume for the second part of the tomato crop cycle (thesis 100-50). Extending the irrigation interval and limiting irrigation volume for the second part of the tomato crop cycle, appeared to be the best management strategy to optimize the yield and nutritional quality of processing tomato.