Construction of mouse ESC line stably expressing Pdx1 by Tet-On system

AIM To construct the mouse embryonic stem cell （ESC） line with stable pancreatic and duodenal homeobox 1 （Pdx1） expression by Tet-On system, which may lay a foundation for further research on the differentiation of Pdx1⁺ definitive endoderm cells into pancreatic cells. METHODS The Pdx1-overexpressing lentiviral vector with green fluorescent protein marker and puromycin resistance was constructed by Tet-On system and was used to infect the mouse ESC. The cells were divided into 3 groups： blank control group （ESC group）, empty lentivirus control group （PDX1^- ESC group） and Pdx1 lentivirus transfection group （PDX1⁺ ESC group）. Flow cytometry was used to detect the transfected cells after screening by doxycycline （DOX）. The function of Tet-On system and the expression of Pdx1 gene were detected. The transfected cells in PDX1^- ESC group and PDX1⁺ ESC group were sorted by flow cytometry, and constructed ESC line with stable expression of Pdx1 and negative control ESC line were verified. RESULTS （1） The positive rates of transfected cells in PDX1^- ESC group and PDX1⁺ ESC group were 90.72% and 94.01% after screening by DOX, respectively. The positive rates of transfected cells in PDX1^- ESC group and PDX1⁺ ESC group was 97.84% and 98.13% after sorting by flow cytometry, respectively. （2） With DOX, green fluorescence was observed in PDX1^- ESC group and PDX1⁺ ESC group. The mRNA and protein expression of Pdx1 was significantly increased in PDX1⁺ ESC group （P<0.05）. Without DOX, no green fluorescence was observed in the cells of the 3 groups, and no significant difference in the mRNA and protein expression of Pdx1 was observed （P>0.05）. （3） After 3 months of cryopreservation, the cell lines still survived in resuscitation culture and were regulated by DOX. CONCLUSION Using Tet-On system, the mouse ESC line with inducible Pdx1 expression were successfully established and could be used as an effective cell model to research the differentiation of Pdx1⁺ definitive endoderm cells into pancreatic cells.     

一种新型二维条码电子耳标在商品猪数字化养殖中的应用

详细阐述了自动识别技术的各种形式。结合当前商品猪养殖的具体情况,本着经济、耐用、实用和对畜产品个体妨碍最小的原则,采用当前比较常用的Grid Matrix码,制作了一种新型二维条码的电子耳标作为畜产品的标签身份号码,为商品猪数字化养殖可溯源管理系统的实现奠定了基础。     

利用CRISPR/Cas9和λ-Red级联技术构建产肠毒素大肠杆菌LT敲除菌株

本研究旨在利用CRISPR/Cas9和λ-Red级联的技术对产肠毒素大肠杆菌（enterotoxigenic Escherichia coli,ETEC）K88的热不稳定性肠毒素（heat-labile toxin,LT）基因进行无痕敲除并获得K88 LT^-缺陷菌株。通过序列比对获取LT两端同源序列,并构建包含LT边界、氯霉素筛选标记、sgRNA和LT同源臂的供体片段;将供体片段转化至ETEC K88,同时分别利用λ-Red同源重组系统和CRISPR/Cas9基因编辑系统,对LT基因进行敲除;通过PCR验证获得了K88 LT^-缺陷菌株,并通过试验测定了敲除菌株的溶血能力和生长曲线。结果显示,λ-Red同源重组系统可成功地将LT基因替换为相应的供体片段,CRISPR/Cas9基因编辑系统可高效地对筛选标记进行删除,最终通过λ-Red和CRISPR/Cas9结合的基因编辑系统可成功对ETEC K88的LT基因进行无痕敲除。体外试验结果表明,K88 LT^-缺陷菌株的溶血能力丧失,并且生长速度比野生型菌株减缓,LT可能和ETEC K88的致病能力和生长性能有关。表明λ-Red和CRISPR/Cas9级联的基因敲除方法可用于LT毒素基因及其他一些大肠杆菌基因的敲除。K88 LT^-缺陷菌株的构建为下一步研究LT毒素的致病机制奠定基础。     

切断深层根对黍子根系及地上部营养生长的影响

采用根管土柱栽培的方法,研究了拔节期和抽穗期切断不同深度(20 cm和40 cm)根系对黍子根系整体及地上部营养生长的影响。结果表明,与不断根相比,在抽穗期和拔节期断掉20 cm或40 cm以下根系,均能导致黍子株高、旗叶叶绿素含量、旗叶SOD与POD活性、单株绿叶面积、总根活力、总根长、总根重及产量明显降低,旗叶M DA含量明显增高。抽穗期断深层根对黍子的影响大于拔节期。但同一生育时期不同深度断根处理间黍子产量差异未达显著水平,表明深层根系(40 cm以下根系)对产量的贡献更大。     

Blood Typing of Asiatic Black Bear (Ursusthibetanus)

In this study,we aimed to determine whether bear had different blood group systems.Whole blood samples were collected from 40 Asiatic Black bears in Fujian province,China.Tube method was used to detect antibodies in plasma,and antibody isotype was determined with 2-mercaptoethanol.Plasma was further analyzed by mass spectrometry.The plasma from four bears had antibodies,possibly IgM isoform,which could agglutinate RBCs from 30 bears.Blood samples from 10 bears were tested by human blood typing reagents.The results showed that 4 bears had blood type like human type O,while 6 bears had like human type B.Plasma protein had extensive homology to serum albumin-like isoform 1 found in giant panda (Ailuropodamelanoleuca).We suggested that Asiatic Black bear might have at least one blood group system with two blood types.If the sick bear needs blood transfusion,a cross-matching test was necessary.Moreover,giant panda might receive blood from Asiatic Black bear in case of emergency.     

土壤质地定名法及吸湿水与土壤粒级含量关系的研究

利用卡庆斯基制、美国制、中国制3种土粒分级标准对张家口葡萄产区的30个采样点进行土壤质地命名,并通过对比分析得出中国制是该区最适的土壤质地分类方法。现对吸湿水含量与土壤各粒级微粒百分含量的关系做了探测性研究,得出吸湿水和土壤中小于0.01 mm的粒径含量之间拟合曲线方程为:Y=20.272e0.207x,可根据此方程应用卡庆斯基制确定土壤质地;吸湿水和土壤中1~0.05 mm0、.05~0.002 mm粒径含量的拟合曲线方程分别为:y=22.018x2-105.323x+156.381,y=-18.304x2+86.495x-49.536,又可根据这2个方程利用美国制确定土壤质地名称。这些为用简易方法鉴定土壤质地类型提供科学依据。     

莫索湾垦区不同土质棉田的土壤水分及群体生理

通过对重壤土和沙壤土两个不同类型的棉田生长状况、植物生理状况和土壤水分状况及土壤温度的研究。结果表明,沙壤土的不保水性和高温使土壤很快变干,造成棉株蒸腾速率下降,气孔阻抗增加,气叶温差变小,形成热害,致使吐絮期提前,影响到了生长状况和经济产量。根据土壤质地造成的灌溉效果的差异,提出在重壤土地采用“大灌量、长周期”,沙壤土地采用“小灌量、短周期”因地配水的灌溉制度,是北疆棉区获得高产应当推行的水分管理技术措施。     

镉对草莓幼苗根尖氧化系统和基因组DNA损伤的影响

采用随机扩增多态性DNA（RAPD）技术和生理生化方法检测镉胁迫对草莓幼苗根尖氧化系统和DNA多态性的影响。结果表明,用5、10和15 mg · L-1镉（CdCl2 · 2.5H2O）处理6 d后,草莓幼苗根伸长及根系中可溶性蛋白质含量均受到抑制,根尖活性氧爆发（超氧阴离子产生速率升高和过氧化氢含量增加）,超氧化物歧化酶、过氧化物酶和过氧化氢酶活性下降,DNA增色效应减少。选用8条寡核苷酸引物（10 bp）对草莓幼苗根尖细胞中基因组DNA进行RAPD扩增,对照组图谱中可分辨出88条RAPD谱带,其分子量为150 ~ 3 500 bp,处理组与对照组RAPD图谱之间存在明显差异,且与镉浓度之间存在剂量效应关系。镉影响草莓幼苗根尖细胞中基因组模板的稳定性,活性氧爆发和DNA交联是根尖DNA损伤的主要原因,利用RAPD技术获得的DNA多态性变化可作为检测草莓根尖DNA损伤的指标。     

系统工程在畜牧业中的应用

本文主要从畜牧业发展战略与规划、遗传育种、营养、养猪、养禽、养羊、养牛等方面阐述了系统工程近二十年来在畜牧业的应用。     

日光温室蔬菜智能化管理系统建模

随着精准农业的发展,智能化管理将在现代农业中广泛应用。针对智能化管理在日光温室蔬菜生产中尚鲜见报道的情况,该研究以日光温室蔬菜为研究对象,通过对日光温室环境特征的分析,设计一种基于无线传感器网络技术的低成本温室监测系统,用于监测温度、湿度、二氧化碳浓度等关键环境参数,并建立日光温室蔬菜系统,通过建模,建立不同子模块,实现蔬菜生产的智能化管理。