Department of Botany and Molecular, Cellular, and Developmental Biology Program, Iowa State University, IA 50011, Ames, USA.
Plant Cell Rep. 1993 May;12(7-8):445-52. doi: 10.1007/BF00234710.
Protoplasts derived from oat (Avena sativa L.) suspension culture cells (7 days after subculturing) were electroporated with plasmid DNA containing the Escherichia coli uidA gene encoding the ß-glucuronidase reporter enzyme. Consistently high enzyme activity was observed with electroporation conditions of 500 μF and 1125 volts/cm. Enzyme activity and mRNA accumulation time courses were determined. The maximum enzyme activity was detected at 24 hours after electroporation, while the maximum mRNA level was detected at 12 hours after electroporation. ß-glucuronidase mRNA was in vitro synthesized with and without a 5' methylated cap and then electroporated into protoplasts. Only capped mRNA produced significant enzyme activity. By electroporating radiolabeled, in vitro synthesized mRNA, the ß-glucuronidase mRNA half-life was estimated to be ∼35 minutes in oat protoplasts.
原生质体来源于燕麦（燕麦 sativa L.）悬浮培养细胞（继代培养后 7 天），用含有编码β-葡萄糖醛酸酶报告酶的大肠杆菌 uidA 基因的质粒 DNA 进行电穿孔。在 500 μF 和 1125 伏特/厘米的电穿孔条件下，观察到一致的高酶活性。测定了酶活性和 mRNA 积累时程。电穿孔后 24 小时检测到最大酶活性，而电穿孔后 12 小时检测到最大 mRNA 水平。β-葡萄糖醛酸酶 mRNA 在有和没有 5' 甲基化帽的情况下进行体外合成，然后电穿孔到原生质体中。只有加帽的 mRNA 产生显著的酶活性。通过电穿孔放射性标记的、体外合成的 mRNA，估计β-葡萄糖醛酸酶 mRNA 在燕麦原生质体中的半衰期约为 35 分钟。
