PEG介导番木瓜叶肉原生质体瞬时表达体系的建立

Establishment of Gene Transient Expression System Mediated by Polyethylene Glycol in Carica papaya L. Mesophyll Protoplasts

本研究对聚乙二醇(polyethylene glycol,PEG)的分子量、浓度及处理时间等三个影响番木瓜叶肉原生质体转化效率的主要因素进行了优化,同时利用绿色荧光蛋白(green fluorescent protein,GFP)表达体系进行了转化表达分析。结果表明:经40%PEG8000溶液处理10 min,可使GFP在原生质体中的平均转化率达到45.26%;沉默抑制载体p Green-Hcpro和GFP瞬时表达载体共转化番木瓜叶肉原生质体,可使GFP在原生质体中的平均转化率进一步提高到68.75%,相比单独转化瞬时表达载体p RNAi-GFP增长了51.90%。PEG介导番木瓜叶肉原生质体瞬时表达体系的建立为番木瓜功能基因组和基因功能研究奠定了基础,可为热带作物的研究提供又一模式作物,推动热带作物在分子及细胞生物学领域的发展。

The polyethylene glycol （PEG） molecular weights, PEG concentration and treatment time were three important factors that influenced the transformation efficiency in transient expression system mediated by PEG in papaya mesophyll protoplasts. A high average transfection efficiency （45.26%） of GFP was obtained when processing samples 10 minutes with 40%PEG8000 solution. In addition, the silence suppression vector of pGreen-Hcpro was successfully constructed. And a higher average transfection efficiency （68.75%） of GFP was obtained when pGreen-Hcpro were co-transformed with the transient expression vector of GFP into the protoplasts. As a result, the transformation efficiency of the co-transformation with GFP and pGreen-Hcpro increased by 51.90% compared with the single transformation. the PEG mediated papaya mesophyll protoplast transient expression system was successfully established, and this study provide the basis for gene silencing suppressor applied in plant bioreactor. And the research may offer a model plant for the study of tropical crops, which would promote the development of tropical crops in molecular and cell biology field.