| Preparation of Primary Cultures of Embryonic Rat Hippocampal and Cerebrocortical Neurons
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Author:
Date:
2017-09-20
[Abstract] This protocol aims at standardizing the procedure to obtain primary cultures of hippocampal and cerebrocortical neurons for in vitro experiments. Cultures should be prepared from cells isolated during embryonic development when neuronal precursor cells are not yet fully differentiated. This helps increasing the quality and quantity of cells, while offering minimal cell death that often occurs during dissociation of differentiated neurons. Cells plated under the appropriate conditions, either in Petri-dishes or in multi-well plates, will develop and establish synaptic contacts over time since the neuronal culture medium provides the nutrients and trophic factors required for differentiation. In this protocol we describe the methodology for the preparation of both cortical and ...
[摘要] 该方案旨在标准化海马和脑皮质神经元原代培养物的体外实验。 培养物应从胚胎发育期间分离的细胞制备，当神经元前体细胞尚未完全分化时。 这有助于提高细胞的质量和数量，同时提供通常在分化的神经元解离期间发生的最小的细胞死亡。 在合适的条件下，在培养皿或多孔板中铺板的细胞将随着时间的推移而发展和建立突触接触，因为神经元培养基提供分化所需的营养和营养因子。 在这个协议中，我们描述了制备皮质和海马神经元培养物的方法。
【背景】本方案描述了使用补充有NeuroCultTM SM1的Neurobasal培养基（Chen等人，2008）的大鼠海马和脑皮层神经元的原代培养物的制备。 NeuroCultTM SM1的组成基于B27补充剂的制剂（Brewer等，1993），但是前者的混合物被发现提高了神经元培养的质量，部分地通过用全转运蛋白替代载脂蛋白转运蛋白 Chen et al。，2008）。 此外，NeuroCultTM SM1的化学成分在原始出版物中有更详细的描述，可以更好地控制实验条件。 用化学确定的培养基制备的神经元培养物的特征在于存在低百分比的星形胶质细胞。 通过添加有丝分裂5-氟-2'-脱氧尿苷的化学抑制剂可以防止维持更长时间的培养物中星形胶质细胞的增殖以允许神经元分化。
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| Quantification of Ralstonia solanacearum Colonization of Potato Germplasm Using Luminescence
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Author:
Date:
2015-05-05
[Abstract] We have developed an unsophisticated, non-disruptive and accurate method for evaluation of pathogen colonization in planta. In this protocol we use a Ralstonia solanacearum (R. solanacearum) UY031 strain genetically modified to constitutively generate light from a synthetic luxCDABE operon stably inserted in its chromosome. This system allows bacterial quantification in a high-throughput manner, avoiding time-consuming and tedious bacterial dilution plating and colony counting. In addition, this system could be especially useful in plant breeding programs to detect bacterial latent growth in symptomless parental lines before their inclusion in long-term disease resistance breeding programs.
[摘要] 我们已经开发了一种不成熟的，非破坏性和准确的方法用于评估植物中的病原体定居。 在该方案中，我们使用经基因修饰以从稳定插入的合成的luxCDABE 操纵子组成性地产生光的（R）solanacearum 其染色体。 该系统允许以高通量方式进行细菌定量，避免耗时和繁琐的细菌稀释电镀和菌落计数。 此外，该系统可以在植物育种程序中特别有用，以在其包括在长期疾病抗性育种程序中之前检测无症状亲本细胞中的细菌潜伏生长。
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