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首页> 外文期刊>Biotechnology Progress >Evaluating Extraction and Storage of a Recombinant Protein Produced in Agroinfiltrated Lettuce
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Evaluating Extraction and Storage of a Recombinant Protein Produced in Agroinfiltrated Lettuce

机译：评价在农用渗滤生菜中产生的重组蛋白的提取和储存

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Extraction and storage of a recombinant protein produced by transient expression following agroinfiltration of lettuce were investigated.Lettuce leaves expressing beta-glucuronidase (GUS)were extracted by homogenization in several buffer combinations,and the yield and stability were assessed.The reducing agent dithiothreitol (DTT)was found to be the most important (significant)component in the extraction buffer.An extraction buffer consisting of 50 mM sodium phosphate at pH 7.0 with 10 mM DTT produced a good yield and stabilized GUS.Leaching of GUS through intact agroinfiltrated lettuce leaves was determined to be infeasible,with a maximum flux of 10 mu g GUS/h/m_2 and recovery of 1.7% of the GUS content in 24 h.Freeze-drying was evaluated as a method to extend the shelf life of the perishable leaf material containing GUS.First-and second-order kinetic models and the Weibull distribution were compared to describe inactivation of GUS in the freeze-dried leaves and leaf extracts.The first-order model best fit the inactivation data.An Arrhenius model fit the first-order inactivation data with respect to temperature with R_2 =1.00.Freeze-drying the lettuce leaves extended the estimated half-life of GUS to 69 days at 21 deg C versus 11 days at 4 deg C for fresh lettuce.
机译：对莴苣进行农杆菌浸润后瞬时表达产生的重组蛋白的提取和贮藏进行了研究。在几种缓冲液组合中均质提取表达β-葡萄糖醛酸苷酶（GUS）的莴苣叶，并评估了产量和稳定性。还原剂二硫苏糖醇（DTT）在提取缓冲液中被发现是最重要的（重要）成分。由pH 7.0的50 mM磷酸钠和10 mM DTT组成的提取缓冲液可产生良好的产量和稳定的GUS。确定不可行，最大通量为10μg GUS / h / m_2，24小时内GUS含量恢复为1.7％。冷冻干燥被认为是延长易腐叶片材料保质期的一种方法GUS。比较一阶和二阶动力学模型以及Weibull分布，以描述冻干的叶片和叶片提取物中GUS的失活。 t阶模型最适合灭活数据.Arrhenius模型适合于R_2 = 1.00的温度的一阶灭活数据。莴苣叶的冷冻干燥将GUS的估计半衰期延长至21摄氏度下的69天相比之下，新鲜生菜在4摄氏度下需要11天。

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