Growth characteristics and transformability of soybean embryogenic cultures

Embryogenic cultures of soybean [Glycine max (L.) Merr. cv. Jack and Asgrow A2872] were established in liquid Finer and Nagasawa medium, maintained by transfer to fresh medium at biweekly intervals, and subjected to microprojectile bombardment over time. Cultures were not amenable to transformation until they were at least 6 months old. Over time, different cell lines of the same genotype acquired very different culture phenotypes. Histological analysis of cell lines differing in transformation ability showed that the most transformable cultures had cytoplasmic-rich cells in the outermost layers of the tissue. In contrast, the outer layers of less transformable cultures contained cells with prominent vacuoles. Although fresh weight accumulation of the cultures was curvilinear during the 2-week subculture period, a burst of mitotic activity was evident shortly after transfer to fresh medium. This activity usually lasted from the 2nd to the 6th day following subculture, and peaked on the 4th day. Tissues at or near this stage always produced more transient expression of a reporter gene than did bombardments at other times. In addition, the cell lines most amenable to transformation also exhibited the highest mitotic index. Thus any treatment to increase the mitotic index, especially when the cell lines are less than 6 months old, may facilitate the transformation of cell lines from which efficient recovery of transgenic plants is still possible.