不同荧光染料对苜蓿原生质体染色效果的研究

Effects of Different Fluorescent Dyes on Staining Alfalfa Protoplast

以紫花苜蓿品种甘农4号(Madicago sativa L.‘Gannong No.4’)愈伤组织酶解的原生质体为材料,采用FDA、罗丹明B、罗丹明6G、吖啶橙和DAPI共5种荧光染料对原生质体进行染色效果比较,以期为原生质体活力测定、细胞核计数及原生质体融合过程的观察提供参考。结果表明:FDA、罗丹明B、罗丹明6G和吖啶橙均可使紫花苜蓿原生质体清晰染色,可用于细胞融合时亲本原生质体的标识。FDA是检测原生质体活力的理想染料;吖啶橙和DAPI可对细胞核进行特异性染色,可用于融合时细胞核的观察和计数,前者还可用于检测融合细胞的活力。通过对异源融合体的鉴别及其活力的测定,为摸索融合条件和融合细胞的培养提供了理论依据。

The effects of five fluorescent dyes(FDA,Rhodamine B,Rhodamine 6G,Acridine Orange and DAPI) on staining protoplasts obtained from callus of Madicago sativa 'Gannong No.4' were investigated in order to test the viability of protoplasts,count nuclei and survey processes of protoplast fusion.Result showed that FDA,Rhodamine B,Rhodamine 6G and Acridine Orange dye and mark protoplasts of alfalfa clearly,which could identify parental protoplasts from fusion clusters.FDA is an optimal dye for testing the viability of protoplasts.Acridine Orange and DAPI can dye specific nucleus of protoplasts,which are then used to count and survey nuclei in cell fusion.Acridine Orange can also test the viability of fusion cells.These studies suggest a basis for exploring cell fusion conditions and culturing fusion cells by identifying heterocaryon and testing their viability.