牛含环形泰勒焦虫血清细胞的生物反应器培养条件的探索

利用生物反应器对牛含环形泰勒焦虫的血清细胞进行了培养条件的探索。结果证明，采用２．５Ｌ培养罐，６片叶轮搅拌，转速１２０ｒ／ｍｉｎ，装量１２００ｍｌ，罐顶一通气孔用沙布棉花封口与外界进行气体交换为最优培养条件。在此状态下，我们对原工艺要求的２０％新生牛血清用量减少至７％，水解乳蛋白也由国产产品替代了部分进口的ＤＩＦＣＯ公司产品，细胞接种密度４．５×１０＾５ｃｅｌｌ／ｍｌ经３６小时培养后平均达２．４×     

碱茅幼穗的组织培养初报

碱茅幼穗在附加有２４－Ｄ１５ｍｇ／Ｌ的ＭＳ培养基中,暗培养条件下两周时形成０５ｃｍ２大小的愈伤组织块,其发生率为４３％。培养３周后形成旺盛生长的肉黄色愈伤组织。愈伤组织的分化是在含ＫＴ１０ｍｇ／Ｌ、ＮＡＡ０５ｍｇ／Ｌ的ＭＳ培养基上２～４周时出现绿色芽点,继续培养１～２周即可得到分化植株,将该植株移至不含激素的ＭＳ或Ｎ６培养基上时,光照条件下形成绿色分化植株。     

奶山羊乳汁中孕酮和17β—雌二醇水平的季节性变化规律

应用ＲＩＡ对１５只西农萨能奶山羊在繁殖季节和非繁殖季节乳汁中的Ｐ４和１７β－Ｅ２水平变化进行研究。结果表明：５～７月份,Ｐ４水平为０９３ｎｇ／ｍｌ±０１０ｎｇ／ｍｌ（０７７～１０９ｎｇ／ｍｌ）,１７β－Ｅ２为５６３３ｐｇ／ｍｌ±４２８ｐｇ／ｍｌ（５０３３～６４２４ｐｇ／ｍｌ）。此时期西农萨能奶山羊处于非繁殖季节。８月中旬以后,试验羊的Ｐ４、１７β－Ｅ２水平均出现与繁殖季节相一致的变化。说明在舍饲条件下,西农萨能奶山羊于８月中旬进入繁殖季节;在首次发情前２ｄ,Ｐ４出现特有的小分泌峰,而在发情期其水平降至很低。配种后,妊娠羊Ｐ４水平一直维持很高,而未孕羊再次出现发情周期的Ｐ４分泌范型,在周期的第１３ｄＰ４水平达峰值（１０９３ｎｇ／ｍｌ±３４５ｎｇ／ｍｌ）。１７β－Ｅ２水平在发情期达到峰值,Ａ组（ｎ＝１２）为８５２７ｐｇ／ｍｌ±１１６１ｐｇ／ｍｌ,Ｂ组（ｎ＝３）为８８０２ｐｇ／ｍｌ±１４３１ｐｇ／ｍｌ。在发情周期的其它时间,１７β－Ｅ２在小范围内波动。     

蜂胶中杀癌物质的分离和定性

早在１９６５年Ｄｅｒｅｖｉｃｉ等人在体外实验证明，蜂胶乙醇水乳剂对癌细胞核和细胞浆有明显的抑制作用。Ｖａｌａｖｉｃｈｙｕ等（１９７５）报道了蜂胶对特定癌细胞培养物的效应。Ｈｌａｄｏｖ等（１９８０）测试了不同溶剂制备的蜂胶提取物对人鼻咽癌和人子宫颈癌细胞系的细胞毒性作用。在规定条件下，５０％的细胞蛋白质合成作用，被半数有效剂量（ＥＤ５０）的蜂胶提取液抑制。他们测得蜂胶乙醚提取液、丁醇提取液及乙酸乙酯提取液的半数有效剂量分别为：３９μｇ／ｍｌ、２６μｇ／ｍｌ和３３μｇ／ｍｌ。Ｓｃｈｅｌｌｅｒ等…     

猪核移植重组胚胎的发育能力

以ＭｃＧｒａｔｈ－Ｓｏｌｔｅｒ（１９８３）提供的去（注）核方法，将猪胚胎的单个卵裂球细胞注入去核的次级成熟卵母细胞，借助电融合的方法（ＡＣ；５Ｖ，１．５ＭＨｚ，１２－１５ｓ；ＤＣ：２ｋＶ／ｃｍ，４０μｓ。电激活／融合液：０．３ｍｏｌ／Ｌ甘露醇＋－．１ｍｍｏｌ／ＬＭｇＳＯ４＋０．１ｍｍｏｌＣａｃＬ２＋７．５ｍｇ／ｍｌＣＣＢ）融入受体胞质构成重级胚，体外培养（培养液为改衣ＴＣＭ－１９９；培养条件为５％     

播娘蒿幼蕾和花序轴的组织培养和再生植株研究

播娘蒿幼小花蕾在附加有１５ｍｇ／ＬＮＡＡ、０５ｍｇ／Ｌ６－ＢＡ和０１ｍｇ／ＬＫＴ的ＭＳ培养基中,培养两周即形成０５ｃｍ２大小的愈伤组织块,出愈率达８３％。继续培养两周转至附加有２ｍｇ／Ｌ６－ＢＡ、０５ｍｇ／ＬＫＴ和０２ｍｇ／ＬＮＡＡ的ＭＳ分化培养基上,２～３周后出现绿色芽点,继续培养２周即可得到分化小芽。播娘蒿花序轴在附加有２～６ｍｇ／Ｌ６－ＢＡ、０５ｍｇ／ＬＮＡＡ的ＭＳ培养基中,培养５～６周也都能分化出芽,但最佳培养基为４ｍｇ／Ｌ６－ＢＡ、０５ｍｇ／ＬＮＡＡ。分化小芽转至附加０５ｍｇ／ＬＮＡＡ或０５ｍｇ／ＬＩＢＡ的１／２ＭＳ生根培养基上培养２～３周均能形成完整植株。     

三至七月龄小尾寒羊营养代谢中甲烷气产量和能值的初步研究

取３月龄断奶小尾寒羊９只随机分为３组（每组１只去势公羊，２只母羊）进行气体能量代谢试验。试验表明，日粮代谢能浓度为２３００～２７００ｋｃａｌ／ｋｇＤＭ、食入代谢能２６００～４８００ｋｃａｌ／ｋｇＤＭ时，甲烷气（ＣＨ４）呼出量２４～３３Ｌ／ｄ，ＣＨ４能（ＣＨ４Ｅ）２２０～３１０ｋｃａｌ／ｄ，甲烷能占食入总能的６％～９％，呼出气体甲烷浓度约为０１０％～０１７％。甲烷能与食入总能回归关系为：３～５月龄：ＣＨ４呼出量（Ｌ／ｄ）＝８７１０６＋０００５２ｘ；ＣＨ４Ｅ（ｋｃａｌ／ｄ）＝７８９７２７＋００５００ｘ；ＣＨ４Ｅ／ＧＥＩ（％）＝１０１５０４－００００８ｘ。５～７月龄：ＣＨ４呼出量（Ｌ／ｄ）＝１１５２１５＋０００４５ｘ；ＣＨ４Ｅ（ｋｃａｌ／ｄ）＝１０８８８３０＋００４２８ｘ；ＣＨ４Ｅ／ＧＥＩ（％）＝１１９４４１－０００１２ｘ。     

H-Y抗血清的制备及检测研究

选用８ ～１２ 周龄纯系的 Ｂ Ａ Ｌ Ｂ／ Ｃ 母鼠, 用同日龄的 Ｂ Ａ Ｌ Ｂ／ Ｃ 公鼠采取皮肤移植和脾细胞注射２ 种方式免疫, 制备 Ｈ Ｙ 抗血清, 经精子微量细胞毒性试验, 筛选抗血清。皮肤移植５ 只小鼠中只有１ 只产生较好的抗血清, 脾细胞注射的４ 只小鼠都能产生较好的抗血清, 产生抗血清的小鼠占５５ ％ 。抗血清与昆明鼠正常８ｃｅｌｌ 早期桑椹胚培养５ ～６ 、１８ 和２４ ｈ 后, 退化率达４３３ ％ , 经 Ｘ２ 检验差异不显著（ Ｐ ＞ ００５） , 符合自然性比例。     

断奶仔猪日粮中添加柠檬酸和氧化锌的试验

试验是研究柠檬酸和氧化锌对仔猪断奶后生长发育的影响。组１为对照组,喂消化能为１３６１ＭＪ／ｋｇ,粗蛋白质为２０％的标准日粮,组２为标准日粮＋１５％柠檬酸,组３为标准日粮２０×１０－４锌,组４为标准日粮＋１５％柠檬酸＋２０×１０－４锌。结果：第１...     

锥-46的耕牛血药浓度反相HPLC测定方法探索

用惠普ＨＰ１０５０四元泵、ＨＰ１０４０Ｍ二极管阵列检测器和色谱柱ＥｃｏｎｏｓｐｈｅｒｅＣ（１８）（２５０×４．６ｍｍ，５μ）探索了锥－４６的耕牛血药浓度测定条件。检测波长２５２±２ｎｍ，参比波长５５０±２ｎｍ。流动相为甲醇：乙腈：重蒸水＝２０：２５：５５（Ｖ／Ｖ），每３００ｍｌ加入二乙胺１０００μｌ，用磷酸调节至ｐＨ３．５，流速０．６ｍｌ／分。血浆中的锥－４６用磷酸酸化至ｐＨ２．４的甲醇溶液提取，平均提取效率达９９．３３±２．２８％。提取物在６０℃下减压浓缩。平均回收率为９９．４６±６．０７％。本法最低检测浓度为０．１μｇ／ｍｌ。在锥－４６浓度为０．１～１０μｇ／ｍｌ范围内线性良好（相关系数ｒ＝０．９９９９，峰面积＝０．６９２８＋１．５８１×锥－４６ｎｇ数）。锥－４６基本上能与血浆中的杂质成分完全分离。已应用于耕牛体锥－４６的药物代谢动力学的研究。     

不同培养基对小鼠黄体细胞体外培养效果的影响

对未性成熟的雌性小鼠注射孕马血清促性腺激素和人绒毛膜促性腺激素,在适当的时间摘取小鼠卵巢进行卵巢黄体细胞的体外培养,培养用液为含10%血清的DMEM和含10%血清的RPMI 1640。结果发现,用前者培养的细胞在形态、规则程度和数量上都优于后者,为小鼠黄体细胞体外培养液的选择和进一步研究提供了参考。     

6株溶磷菌和4株固氮菌混合培养条件的研究

对6株溶磷菌和4株固氮菌混合培养,选择最适培养条件即温度,pH和盐分,为确定促生菌的最佳发酵条件和生态适应性评价提供依据。结果表明：溶磷菌和固氮菌单独培养或混合培养最适温度在25-35℃,混合培养较单独培养生长良好;溶磷菌和固氮菌单独培养或混合培养最适pH为6．5～7．5,在pH7．5～8．5偏碱性环境下,混合培养生长...     

类植物乳杆菌L-ZS9培养基及培养条件的优化

类植物乳杆菌L-ZS9分离自发酵肉类食品,作为益生菌制剂和发酵剂在乳品行业具有良好的应用潜力。为实现类植物乳杆菌L-ZS9的高密度培养,以MRS培养基为基础进行培养基的优化,并优化培养条件。以单因素试验初步确定培养基中所用的碳源和氮源,通过Plackett-Burman试验确定出培养基成分与培养条件中影响较大的因素,经最陡爬坡试验确定各因素的水平,再通过响应面旋转中心组合试验拟合出菌体密度与因素水平间的方程,从而确定优化后的培养条件。结果表明：培养基中所用的碳源为蔗糖,氮源为酵母粉;经过验证实验得到的优化     

Establishment of optimal conditions for long-term culture of erythrocytic stages of Theileria uilenbergi

OBJECTIVE: To establish optimal conditions for long-term culture of the erythrocytic stage of Theileria uilenbergi. SAMPLE POPULATION: Red blood cells from 3 splenectomized sheep experimentally infected with a blood stabilate of T uilenbergi. PROCEDURES: Cultures of T uilenbergi were initiated by use of blood from experimentally infected sheep collected when parasites were detected in Giemsa-stained thin blood smears. Different culture conditions were tested to optimize in vitro growth of the organisms. Subcultures were performed at a ratio of 1:2, 1:4, and 1:8 when the percentage of parasitized erythrocytes (PPE) was at least 1% or when the initial PPE was doubled. RESULTS: The optimal culture medium was HL-1 medium (a complete chemically defined medium) supplemented with 20% sheep serum and 0.75% chemically defined lipids. Optimal culture conditions included incubation in a humidified 2% O(2), 5% CO(2), and 93% N(2) atmosphere at 37 degrees C. Cultures of the merozoite stage of the parasite were continuously propagated in vitro for > 1 year. The PPE reached values of up to 3%. CONCLUSIONS AND CLINICAL RELEVANCE: Optimization of culture conditions to reach a high PPE seems worthwhile. The continuous propagation of T uilenbergi in culture allows the production of parasite material without infecting animals and provides a continuous laboratory source of parasites for further studies.     

犬肾（DK）传代细胞中霉形体的分离与鉴定

取经电镜观察证实有霉形体污染的ＤＫ传代细胞，分别接种于霉形体检验用液体、半流体和固体培养基，证明为霉形体混合感染，即该培养物在液体培养基和半流体培养基上，同时显示出水解精氨酸和发酵葡萄糖２种特性，前者使培养基ｐＨ上升，后者使ｐＨ下降。通过反复挑选单个菌落进行克隆传代和有目的地加入单一种属的霉形体抗血清的方法，将混合的２株霉形体纯化分开。经鉴定，一株为精氨酸霉形体，一株为莱氏无（需）胆甾醇霉形体。     

青藏高原高寒草甸土壤N_2O排放通量对温度和湿度的响应

试验采用室内培养的方法研究了青藏高原矮嵩草草甸土壤氧化亚氮(N2O)排放通量对土壤温度和湿度的响应过程。结果表明:随培养温度增加,培养初期矮嵩草草甸土壤N2O排放速率逐渐降低(P0.05),最高排放速率为4.75±0.24g/(kg·h),最低排放速率仅为2.92±0.19g/(kg·h);而经过7d培养,高寒草甸土壤N2O排放速率先降低,而且转变为弱汇,而后升高,30℃时最高,为0.67±0.06g/(kg·h),各处理均显著低于短期培养时排放速率(P0.05);随土壤湿度增加,短期培养矮嵩草草甸土壤N2O排放速率先降低随后升高,在土壤湿度为60%时排放速率最高,为(3.62±0.38)g/(kg·h),而在75%时排放速率最低,为(3.38±0.25)g/(kg·h);随着培养时间延长,高寒草甸土壤N2O排放速率逐渐降低,最高排放速率为(0.55±0.32)g/(kg·h);土壤湿度分别为45%和温度为20℃、以及60%和30℃时,1d和7d培养矮嵩草草甸土壤N2O排放速率最高;而在土壤湿度为45%和30℃、以及75%和30℃时,1d和7d培养矮嵩草草甸土壤N2O排放速率最低,后者呈现吸收现象,吸收速率约为-1.84g/(kg·h)。     

鸡源凝结芽孢杆菌的分离鉴定及耐受性分析

【目的】试验旨在寻找1株性能稳定、环境耐受性强的凝结芽孢杆菌菌株,以在实际生产中发挥其替抗功能。【方法】本研究采集160日龄健康海兰褐蛋鸡的粪便样本,进行细菌分离纯化,并对分离菌进行形态学特征观察、生理生化鉴定、16S rDNA测序分析。通过绘制该菌的生长动力曲线确定其生长规律;利用耐酸、耐胆盐试验分析该菌株的耐受性;通过单因素试验对分离菌株的培养条件进行优化。【结果】分离菌在普通培养基上长出表面粗糙、不规则的灰白色菌落,镜检为杆状、芽孢端生的革兰氏阳性菌。结合生化试验及16S rDNA基因序列分析综合鉴定该菌株为凝结芽孢杆菌,命名为NJ001。生长动力曲线表明,7～24 h为该菌株的对数生长期,24 h活菌数达到顶峰,凝结芽孢杆菌NJ001的最佳培养时间为24 h;耐受性试验结果显示,该菌株pH 3.0温育2 h存活率为59.5%,pH为4.0时存活率达到87.6%以上,对酸有较强耐受力;0.3%胆盐处理2 h存活率为54.0%,胆盐浓度升至0.5%时存活率仍有50%以上,该菌对胆盐有较强的耐受性。单因素试验结果发现,该菌的最佳培养温度为40℃,接种量2%,初始pH 7.0,摇床转速...     

布氏锥虫伊氏亚种体外培养的研究——Ⅲ.集落的形成及其生物学意义

以无细胞培养系统培养布氏锥虫伊氏亚种(Trypanosma brucei subsp.evansi,简称伊氏锥虫),可见其有规律地形成集落.在每日混匀换液后8h开始虫体向局部聚集,形成集落;随着聚集过程的进展,集落范围缩小,虫密度增高,与周围境界愈加明显.12h集落大小趋于稳定,而虫密度仍持续上升至18h其表现为集落内虫体向多层立体分布发展.虫体不附着于培养器皿,运动活泼.在集落虫密度不断增高的同时,集落内单个虫体逐渐减少,而由分裂态虫体形成的小虫簇相应增多.集落实际上是旺盛分裂增殖的虫体群落.从培养8h起,每2h把培养物混匀1次,其前12h虫密度增长情况与始终静置培养的培养物基本一致,但12h后不再增殖,而后者则持续增殖至18h致使前者的最高虫密度仅为后者的约3/4.结果说明,集落的形成和保持有助于生长繁殖,是伊氏锥虫固有的生物学特性.本文还讨论了其形成机制和生物学意义.     

植物乳杆菌(Lactobacillus plantarum)8-6产细菌素发酵条件的优化

对植物乳杆菌(Lactobacillus plantarum)8-6产细菌素的发酵条件进行了优化,分别研究了培养时间、温度、接种量、培养基起始pH值、培养基碳源、氮源等因素对细菌素产生的影响,通过单因素水平试验和正交试验,确定产细菌素的最佳培养基组合和最佳发酵条件为葡萄糖3%,胰蛋白胨2%,蛋白胨1%,酵母膏1%,硫酸镁0.058%,吐温-80 0.2%,30℃培养24h,培养基起始pH值为6.5,接种量2%。乳杆菌8-6优化后效价为1825.56IU/mL,比优化前提高了373.15%。     

In vitro development of mouse pronuclear embryos to blastocysts in sequential media with and without co-culture of autologous cumulus cells

The objective of this study was to use mouse embryos as a model system to investigate the effect of co-culture of cumulus cells in Sydney IVF sequential media (Cook) on embryo development, based on the hypothesis that feeder cells in co-culture with a sequential medium could work synergistically to further improve in vitro culture conditions for mammalian preimplantation embryos. The culture systems described here were evaluated by the ability to consistently produce high blastocyst formation rates and high cell number per blastocyst. The role of embryo-to-cell contact was assessed by using Transwell inserts with transparent microporous membranes. Pronuclear embryos of ICR mice were cultured to blastocysts in Cook sequential media, with and without mouse primary cultures of cumulus cells, and with or without inserts. Blastocyst formation rates and cell numbers of in vitro developing embryos in the different culture systems were compared to each other, and to in vivo derived blastocysts. Blastocyst formation rates for Cook medium only was 27.8% (without inserts) and 32.9% (with inserts), whereas Cook-Cumulus cells in identical culture systems was significantly higher at 45.8% (without inserts, P<0.05) and 55.6% (with inserts, P<0.05). When the embryos are suspended above the bottom of the well, for Cook medium significantly lower blastocyst formation rates were observed at 4.2% compared to Cook-Cumulus cells at 17.5% (P<0.05). Mean cell numbers of blastocysts obtained in all co-culture systems were significantly higher (P<0.05) compared to those developing in culture medium only. Although the putative mechanism is as yet unexplained, the improved blastocyst formation rates and cell numbers in co-culture when there is direct contact between the embryo and the cell monolayer suggest that the close proximity between the feeder cells and embryos is in part responsible for these effects.