Makonkawkeyoon S, Smitamana P, Hirunpetcharat C, Maneekarn N
Department of Microbiology, Faculty of Medicine, Chiang Mai University, Thailand.
Experientia. 1995 Jan 15;51(1):19-25.
Mouse-tobacco hybrid calli, and complete plants producing mouse gamma-3 heavy and lambda light chains, have been generated by somatic cell fusions of mouse spleen cells and tobacco mesophyll protoplasts. Both gamma 3 and lambda chains were detected in hybrid calli and complete plants by enzyme-linked immunosorbent assay, immunofluorescent staining, and Western blotting. When cellular DNA from hybrid tobacco mesophyll protoplasts was amplified by the polymerase chain reaction (PCR) using two pairs of gamma 3 chain DNA primers and one pair of lambda chain DNA primers, the PCR products contained gamma 3 and lambda chain DNAs, which could be detected by southern blotting and DNA hybridization, using specific synthetic oligonucleotide probes for gamma 3 and lambda respectively. In situ hybridization of hybrid tobacco mesophyll protoplasts with specific recombinant DNA probes of gamma 3 and lambda chains showed the presence of gamma 3 and lambda chain DNAs in the hybrid protoplasts.
通过小鼠脾细胞与烟草叶肉原生质体的体细胞融合,已获得小鼠 - 烟草杂种愈伤组织以及产生小鼠γ-3重链和λ轻链的完整植株。通过酶联免疫吸附测定、免疫荧光染色和蛋白质印迹法在杂种愈伤组织和完整植株中检测到了γ3和λ链。当使用两对γ3链DNA引物和一对λ链DNA引物通过聚合酶链反应(PCR)扩增杂种烟草叶肉原生质体的细胞DNA时,PCR产物包含γ3和λ链DNA,使用分别针对γ3和λ的特异性合成寡核苷酸探针,通过Southern印迹和DNA杂交可检测到这些产物。用γ3和λ链的特异性重组DNA探针与杂种烟草叶肉原生质体进行原位杂交,结果显示杂种原生质体中存在γ3和λ链DNA。