百合原生质体分离培养和瞬时转化  

Isolation,cultivation,and transient transformation of lily protoplasts

作　　者：何珊珊 李宏宇[1,2] 马月 孙红梅 HE Shanshan;LI Hongyu;MA Yue;SUN Hongmei(Key Laboratory of Protected Horticulture of Ministry of Education/National and Local Joint Engineering Research Center of Northern Horticultural Facilities Design and Application Technology,College of Horticulture,Shenyang Agricultural University,Shenyang 110866,Liaoning,China;College of Life Science and Bioengineering,Shenyang University,Shenyang 110044,Liaoning,China)

机构地区：[1]沈阳农业大学园艺学院,设施园艺教育部重点实验室/北方园艺设施设计与应用技术国家地方联合工程研究中心,辽宁沈阳110866 [2]沈阳大学生命科学与工程学院,辽宁沈阳110044

出　　处：《浙江大学学报(农业与生命科学版)》2025年第1期67-79,共13页Journal of Zhejiang University:Agriculture and Life Sciences

基　　金：辽宁省种质创新藏粮于技专项计划项目(2023JH1/10200010);国家现代农业产业技术体系建设专项(CARS-23)。

摘　　要：原生质体是遗传转化和基因功能验证的重要受体。百合(Lilium)是世界上重要的观赏、食用和药用植物,其原生质体分离培养体系仍不完善。本文以细叶百合(Lilium pumilum)和新铁炮百合(Lilium formosanum×Lilium longiflorum var. scabrum)为材料,对百合原生质体分离培养和瞬时转化进行了深入探索。结果表明:百合叶片和胚性愈伤组织均是分离原生质体的优良材料。试管苗叶片原生质体的最佳分离液为细胞原生质体清洗液(cell protoplast wash medium, CPW)+1.0%~2.0%纤维素酶RS+0.5%离析酶R-10+0.10%果胶酶Y-23+12~14 g/L D-甘露醇。胚性愈伤组织原生质体分离的最佳分离液为CPW+2.0%纤维素酶RS+0.60%果胶酶Y-23+12~14 g/L D-甘露醇。叶片原生质体是瞬时转化的优良受体,瞬时转效率为34.0%~36.7%。新铁炮百合胚性愈伤组织原生质体具有很强的分裂能力,在MS(Murashige-Skoog,含206.25 mg/L NH_(4)NO_(3))+60 g/L葡萄糖的固-液双层培养基中及培养密度为2×10^(5)个/mL条件下,培养70 d可形成愈伤组织。本研究为百合细胞工程和分子育种奠定了基础。Protoplasts are important receptors for genetic transformation and gene function verification.Lily is an important ornamental,edible,and medicinal plant worldwide.The isolation and cultivation system for lily protoplasts is still incomplete.In this study,taking Lilium pumilum and Lilium formosanum×Lilium longiflorum var.scabrum as materials,the isolation,cultivation,and transient transformation of lily protoplasts were studied.The results revealed that lily leaves and embryogenic callus tissues were both excellent materials for isolating protoplasts.The optimal solution for isolating protoplasts from sterile plantlet leaves was cell protoplast wash medium(CPW)+1.0%-2.0%cellulase RS+0.5%macerozyme R-10+0.10%pectinase Y-23+12-14 g/L D-mannitol.The optimal solution for isolating protoplasts from embryogenic callus tissues was CPW+2.0%cellulase RS+0.60%pectinase Y-23+12-14 g/L D-mannitol.Protoplasts isolated from leaves were excellent receptors for transient transformation,with a transformation efficiency of 34.0%-36.7%.Protoplasts isolated from embryogenic callus tissues of Lilium formosanum×Lilium longiflorum var.scabrum had strong division ability.In a solid-liquid double-layer culture medium with MS(Murashige-Skoog,containing 206.25 mg/L NH_(4)NO_(3))+60 g/L glucose and a cultivation density of 2×10^(5) cells/mL,callus tissues were formed after 70 days of cultivation.This study lays the foundation for lily cell engineering and molecular breeding.

关 键 词：细叶百合 新铁炮百合 原生质体培养 瞬时转化 

分 类 号：S682.29[农业科学—观赏园艺]