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枯草芽孢杆菌突变株IA857高效转化方法的研究 被引量:1
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作者 牛福星 于晶晶 +3 位作者 汤宏赤 刘金平 滕昆 韦宇拓 《广西科学》 CAS 2014年第2期119-123,共5页
【目的】研究比较枯草芽孢杆菌突变株IA857高效转化的最优方法。【方法】以质粒pHCMC04(大小为8089bp)转化突变型枯草芽孢杆菌IA857,分别用电转化法、原生质体法、电击法诱导的原生质体法、Spizizen低盐环境感受态转化法(简称S法)、低... 【目的】研究比较枯草芽孢杆菌突变株IA857高效转化的最优方法。【方法】以质粒pHCMC04(大小为8089bp)转化突变型枯草芽孢杆菌IA857,分别用电转化法、原生质体法、电击法诱导的原生质体法、Spizizen低盐环境感受态转化法(简称S法)、低盐环境形成饥饿感受态法及其改良方案在其它突变菌株的试验方法(简称C法)进行实验,通过比较优化得出最优转化方法。【结果】采用S法转化枯草芽孢杆菌突变株IA857,质粒加入量为70ng时,转化后直接摇床培养1h转化率最高达到1.2×104 CFU/μg DNA,可以满足枯草芽孢杆菌高效转化的要求。【结论】得出了枯草芽孢杆菌突变株IA857最优转化方法并提出转化枯草芽孢杆菌的基本框架。 展开更多
关键词 枯草芽孢杆菌 转化 电击法 原生质体法 低盐环境 感受态
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实验条件对心叶烟接种TMV后枯斑产生时间及数目影响 被引量:1
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作者 谭翔 《科技信息》 2013年第17期189-190,共2页
在光照60%、相对湿度70%下,测试温度、TMV浓度对心叶烟出斑数和出斑时间的影响。同时,用原生质体法,建立原生质体中复制的TMV的最佳接种浓度。得出结论:心叶烟温度在26-30℃,TMV浓度在24-29μg/ml(质量浓度)时,每半片叶出斑约60个,出斑... 在光照60%、相对湿度70%下,测试温度、TMV浓度对心叶烟出斑数和出斑时间的影响。同时,用原生质体法,建立原生质体中复制的TMV的最佳接种浓度。得出结论:心叶烟温度在26-30℃,TMV浓度在24-29μg/ml(质量浓度)时,每半片叶出斑约60个,出斑时间约70小时,为半叶枯斑法的最佳条件。 展开更多
关键词 TMV 心叶烟 原生质体法 最佳条件
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Flow Cytometric Evidence for Hydroxyl Radical-induced Apoptosis in Tobacco Protoplasts 被引量:4
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作者 雷晓勇 廖旭东 +1 位作者 张贵友 戴尧仁 《Acta Botanica Sinica》 CSCD 2003年第8期944-948,共5页
Protoplasts prepared from tobacco (Nicotiana tabacum L., cultivar BY-2) suspension cells have similar morphological characteristics to those in animal cells. The hallmarks of apoptosis such as condensation and periphe... Protoplasts prepared from tobacco (Nicotiana tabacum L., cultivar BY-2) suspension cells have similar morphological characteristics to those in animal cells. The hallmarks of apoptosis such as condensation and peripheral distribution of nuclei, TUNEL positive reaction, and DNA ladders were observed when tobacco protoplasts were treated with the hydroxyl radical generating system (1.0 mmol/L FeSO4/0.5 mmol/L H2O2). In animals, the loss of transmembrane potential (DeltaPsi(m)) and the exposure of phospholipid phosphatidylserine (PS) are believed to be the main apoptosis events. To test whether these significant processes take place in plants, flow cytometry was used to detect annexin V binding and changes in DeltaPsi(m). Results showed that the PS turned out from inner membrane and DeltaPsi(m) gradually decreased during the apoptosis. All these apoptotic characteristics proved that hydroxyl radicals can cause typical programmed cell death (PCD) in tobacco protoplasts and this design can be served as an effective experiment system to explore the mechanism of plant apoptosis. 展开更多
关键词 tobacco protoplasts flow cytometry APOPTOSIS programmed cell death (PCD) hydroxyl radicals
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PEG-mediated Gene Transfer into Orychophragmus Violaceus Cotyledon Protoplast and Regeneration of Transgenic Plants
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作者 周冀明 卫志明 +1 位作者 刘世贵 罗鹏 《Developmental and Reproductive Biology》 1994年第2期55-63,T001,共10页
Transgenic plants were obtained by PEG-mediated tranfer of foreign gene into cotyledon protoplasts of Orychophragums violaceus. Systematic study was carricd out on PEG-mediatcd transformation of cotyledon protoplast u... Transgenic plants were obtained by PEG-mediated tranfer of foreign gene into cotyledon protoplasts of Orychophragums violaceus. Systematic study was carricd out on PEG-mediatcd transformation of cotyledon protoplast using transient expression system, which showed 25-30 μg of pasmid, 15% PEG and a pH value of 8.0 as the optimal parameters contributing to the highest expression level. Using these parameters, cotyledon protoplasts were isolated, treated with bacterial plasmid DNA (pBI222 with HPT as selective marker) and PEG, and cultured at a density of 5×10 4/ml.After 10-15 days,they were selected by adding 25 μg/ml hygromycine. One month later, a few calli were observed, which were then transferred onto a solid medium with 50-100 μg/ml hygromycine for proliferation. Later they were transferred successively onto differentiation and rooting media and finally hygromycineresistant whole plants were obtaincd. The plants grew well in pots and a regeneration rate of 5 ×10(-5) was achieved. Then,excised leaves of the transgenic plants were used as explants for Southern blot analysis, which confirmed the stable integration of HPT gene into the chromosomal genome of Orychophragmus violaceus The transformation frequency was 10-5. 展开更多
关键词 Orychophragmus violaceus COTYLEDON Protoplast culture PEG transformation Transgenic plant.
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