嗜水气单胞菌外膜蛋白（OMP）的原核表达及其对BALB/c小鼠的免疫保护研究

为确定嗜水气单胞菌(Aeromonas hydrophila,Ah)AS1.927株外膜蛋白(OMP)的原核表达产物是否具有抗原性及其对小鼠的免疫保护力.本研究根据GenBank上嗜水气单胞菌外膜蛋白ompA基因序列(GenBank登录号:AF146597)设计1对引物,克隆出ompA全长基因1020 bp(GenBank登录号:EU309491),经Sal Ⅰ和Xhol Ⅰ双酶切后连接pET-30a(+),转化大肠杆菌(Escherichia cDli)BL21,IPTG诱导表达,并进行SDS-PAGE及Westernblot分析鉴定.结果显示重组基因工程菌E.coli[pET-30a-ompA]表达的融合蛋白分子量约为40.7kD,可以被鼠抗嗜水气单胞菌外膜蛋白抗血清识别.将重组基因工程菌Ecoli[pET-30a-om-pA]口服免疫BALB/c小鼠(Mus mussulus);末次免疫1周后用放射免疫分析小鼠肠粘膜分泌型免疫球蛋白A(sIgA)水平,检测结果揭示,该重组基因工程菌能提高sIgA的分泌(P<0.05);同时在小鼠血清中测出特异性抗体IgG(P<0.05).末次免疫2周后用100 LD50的Ah AS1.927(3.3×105> cfu/mL)腹腔注射攻击小鼠,口服重组菌对小鼠的相对免疫保护力(relative percent survival,RPS)为75%,表明重组基因工程菌可诱导小鼠对AhAS1.927产生一定的免疫保护作用.     

多重实时RT-PCR快速检测新型甲型H1N1流感病毒

从GenBank中获取新型甲型H1N1流感病毒(2009年流行株)HA和NA基因序列,通过多重序列比对之后,设计合成H1和N1基因特异性的引物和探针,用来建立新的甲型H1N1亚型流感病毒的多重实时RT-PCR检测方法.合成的两条荧光探针分别标记FAM和CY5荧光报告基团,荧光淬灭均使用BHQ基团.多重实时RT-PCR实验在ABl7500实时PCR仪上进行,经过40个循环的扩增之后,阳性对照出现标准的S型曲线,并且具有良好的特异性,可以很好地将新的甲型H1N1与传统的甲型H1N1以及H3N2、H5N1、H6N2和H9N2等流感病毒区分.多重实时RT-PCR可以检测到10个拷贝的模板,灵敏度接近检测方法的极限.检测时间短,从加样到反应结束只需要90 min.在对243例发热病人临床样品检测过程中发现7例阳性,1351份猪临床样品检测中未发现阳性,该检测结果和采用中国检验检疫科学研究院研制的试剂盒获得的检测结果一致.本研究所建立的快速、准确和高敏感性的多重实时RT-PCR方法非常适用于新型甲型H1N1流感病毒的实验室筛查.     

重组MBP-GnRH6融合蛋白的表达与鉴定

将人工合成促性腺激素释放激素(GnRH)六聚体基因片段连接至pMAL-c4x载体,获得的pMAL-c4x-GnRH6重组质粒转化到大肠杆菌TB1中进行表达.采用SDS-PAGE,Western blot进行鉴定,并通过动物实验对表达产物的免疫原性进行检测.获得了分子量大小约为50 kD的麦芽糖结合蛋白-GnRH六聚体(MBP-GnRH6),且融合蛋白与GnRH抗体的反应性较好,并能使睾丸萎缩、变性.表明重组MBP-GnRH6融合蛋白具有较好的反应原性和免疫原性,可用于小鼠的免疫去势.     

口服急性毒性试验中Cry1C蛋白对鼠肠道菌群的影响

利用变性梯度凝胶电泳(DGGE)分析不同ICR(institute of cancer researcch)小鼠在口服急性毒性试验中摄入Cry1C蛋白前后其肠道菌群的变化,进而对该抗虫蛋白的食用安全性进行较为深入的研究.DGGE图谱经UPGAMA聚类分析显示:小鼠间菌群结构存在较明显的个体差异;实验期间,对照组小鼠灌胃前后其菌群基本保持稳定;实验组小鼠试验前后肠道菌群变化差异较对照组明显.随着灌胃的停止其变化差异在经历过高峰后会随着停喂时间的延长而减小,且菌相有逐渐恢复灌胃前结构的趋势.从肠道菌相的角度分析可知:该抗虫Cry1C蛋白对小鼠是基本安全的.     

聚乙二醇（PEG6000）模拟干旱胁迫抑制矮沙冬青种子的萌发

本研究以新疆特有濒危保护植物矮沙冬青（Ammmopiptanthus nanus（M．Pop．）Chengf）的种子为材料,用不同渗透势浓度的聚乙二醇（PEG6000）模拟干旱胁迫,探讨干旱胁迫对矮沙冬青种子发芽率、平均发芽速度、胚轴和胚根长度及发芽指数、活力指数的影响。结果表明,不同浓度PEG胁迫处理均降低了种子的发芽率,延缓了矮沙冬青种了萌发进程;种子的发芽率、发芽指数和活力指数均随胁迫强度的增加呈明显下降趋势。当-1．20MPa的PEG胁迫处理的种子在试验结束时仍未能萌发,表明-1．20MPa是矮沙冬青种子萌发的临界水势。PEG模拟十旱胁迫中,当PEG处理为-0．2MPa时,虽然最终发芽率与对照一样,但其胚根、胚轴的长度都比对照短,说明矮沙冬青胚根、胚轴的生长比发芽率对干旱胁迫更敏感。干旱胁迫可能是导致矮沙冬青种群天然更新能力弱的原因之一。本研究将为矮沙冬青种质资源的保护和种群的恢复提供科学依据。     

Fate of prions in soil: Degradation of recombinant prion in aqueous extracts from soil and casts of two earthworm species

Prions represent the active agent in transmissible spongiform encephalopathy (TSE) diseases and can remain infective to mammals even after prolonged periods in soil. The influence of mesofauna on prion dispersal and degradation in soil, however, remains unknown. In this study the effect of earthworms on the retention/dissemination of TSEs in soil was evaluated using a model recombinant prion protein (recPrP) and aqueous extracts from soil and fresh casts of two earthworm species, Lumbricus terrestris and Aporrectodea caliginosa. Our results showed that earthworm gut-derived enzymes did not enhance the degradation of recPrP in comparison to soil, even though non-prion related proteolytic activity was higher in fresh worm excrements than in soil samples. Complete degradation of recPrP occurred in the aqueous extracts from all samples within up to 6 days at +15 °C. The proteolytic enzymes responsible for degrading recPrP were inhibited by aprotinin and leupeptin and studies in pure cultures suggested these were most probably of soil microbial origin.     

Relationship between MAP-2, nestin and development of muscular layer of human embryonic esophagus

AIM: To explore the relationship between microtubule-associated protein 2 (MAP-2), nestin and development of esophageal muscular layer in human embryos. METHODS: At the second to fourth month of gestation, the protein expression of MAP-2 and nestin was observed using the PV method of immunohistochemistry in the human embryonic esophageal tissues. RESULTS: At the second month, the third month and the fourth month of gestation, MAP-2 positive expression in the neural cells of the intermuscular nerve plexus, and negative expression in the muscle cells of human embryonic developing esophageal muscle were observed. Nestin positive expression was found in the muscle cells at the second month to the third month of gestation. However, no nestin expression in the muscle cells of the esophageal muscle at the fourth month of gestation was observed. At the second month to the fourth month of gestation, nestin showed negative or weak positive expression in the neural cells of the intermuscular nerve plexus of human embryonic esophageal tissues. CONCLUSION: MAP-2 and nestin might regulate the formation of esophageal muscular layer in human embryos.     

籽粒苋鲜茎叶蛋白质含量及叶蛋白提取效率研究

对Ｋ１１２、Ｒ１０４等８个籽粒苋品种分不同时期进行了鲜茎叶蛋白含量分析和叶蛋白提取试验研究。结果表明，籽粒苋属于高蛋白质含量作物，全生育期叶蛋白含量平均达２８．７６％，采穗后茎叶平均蛋白质含量仍可达１１．１５％，其最佳利用时期为现蕾期至开花期，其中Ｒ１０４、Ｒ１０１１、Ｋ５７４蛋白质含量较高。     

人参设施栽培研究进展

该文归纳总结了设施内部环境对人参生长发育的影响,并对国内外利用设施栽培人参的相关试验结果及文献资料进行汇总,从设施内部环境、设施的结构与材料、设施内环控设备方面阐述了各种因素对人参生长发育的影响,为利用设施栽培人参提供可靠理论依据。