人睫状神经营养因子的原核表达,纯化及其生物效应

人睫状神经营养因子（ｈＣＮＴＦ）克隆入ｐＢＶ２２０中，在ＤＨ５α菌株中表达，重组蛋白以包含体的形式存在，表达量为菌体总蛋白的５０％左右。经比较发现用２ｍｏｌ／Ｌ脲洗涤包含体可溶解大量可溶性细菌蛋白，且包含体损失较小。在高浓度变性剂条件下进行ｓｅｐｈａｒｃｙｌＳ－２００凝胶过滤，解决了纯化中ｈＣＮＴＦ易聚合的问题，在低浓度变性剂条件下进行ＤＥＡＥ离子交换，有利于蛋白活性的保持。经两步纯化后得到均一性ｈＣＮＴＦ，纯度达９５％以上。在自然状态下使ｈＣＮＴＦ复性。纯化复性后的ｈＣＮＴＦ对无血清培养的鸡胚背根节神经元和脊髓腹角运动神经元有明显的维持存活和促进生长发育的生物效应。     

MacStylet, software to analyse electrical penetration graph data on the Macintosh

Conclusion Since the EPG method is increasingly utilized in the investigation of plant-Homoptera interactions, this software has been developed to enable fast processing of abundant data. The objective seems to have been achieved and, with a little practice, a 2-hour experiment may be analysed in about 10–15 minutes. Mac-Stylet is stand-alone shareware, freely distributed to all persons interested (request to G. Febvay, email: febvay@jouy.inra.fr).     

Essential role for induced Ca2+ influx followed by [Ca2+]i rise in maintaining viability of yeast cells late in the mating pheromone response pathway. A study of [Ca2+]i in single Saccharomyces cerevisiae cells with imaging of fura-2

We established an experimental system for measuring the cytosolic-free Ca2+ concentration ([Ca2+]i) in individual Saccharomyces cerevisiae cells using fura-2 as a Ca2(+)-specific probe in conjunction with digital image processing and examined changes in [Ca2+]i in response to alpha-factor in single cells of a mating type. The addition of alpha-factor to a cells raised [Ca2+]i to several hundred nanomolar in the cells from a basal level of approximately 100 nM, simultaneous with the induction of Ca2+ influx. When the cells were incubated with alpha-factor in a Ca2(+)-deficient medium, Ca2+ influx was greatly reduced, and the rise in [Ca2+]i was not detected. This indicates that the alpha-factor-induced rise in [Ca2+]i is generated by Ca2+ influx through the plasma membrane and not by release from internal stores. In the Ca2(+)-deficient medium, a cells died specifically after they had changed into cells with one projection on the cell surface. This indicates that the rise in [Ca2+]i is essential for the late response to alpha-factor. The duration of Ca2+ requirement for maintaining viability was limited to this stage, and the earlier and later stages were not affected by Ca2+ deprivation. Mating between a and alpha mating type cells was impaired in this medium due to cell death at and before the stage of conjugation. These findings are the first evidence for an essential role for mobilized Ca2+ in the yeast life cycle.     

The dynamics of the subtalar joint in sudden inversion of the foot

The human subtalar joint was modelled as a quasi-linear second-order underdamped system to simulate sudden inversion motion of the foot relative to the shank. The model was fed with experimental data obtained from six subjects on a specially constructed apparatus. A total of 35 deg inversion was produced on the tested leg rapidly enough (lasting less than 40 ms) in order to ensure that the protective muscles are not activated. The parameters of the joint were evaluated and the following ranges were obtained at 35 deg inversion: elastic stiffness 14-52 Nm rad-1, damping coefficient 1.4-2.9 Nms rad-1, and natural frequency 78-125 Hz. The effects on the test parameters of weight bearing amount, foot dominance, and protective footwear were studied on one subject.