Wetterauer B, Morandini P, Hribar I, Murgia-Morandini I, Hamker U, Singleton C, Macwilliams H K
Zoologisches Institut, Ludwig-Maximilians-Universität, Luisenstrasse 14, Munich, 80333, Germany.
Plasmid. 1996 Nov;36(3):169-81. doi: 10.1006/plas.1996.0044.
Almost all methods for transformation of the social ameba Dictyostelium discoideum rely on axenic growth, that is, growth in a synthetic medium, for at least part of the procedure. Axenic growth requires several mutations. Here we describe a procedure that can be used to transform wild-type strains which are able to grow only on the natural food source, bacteria. The method relies on a new selection cassette driven by the V18 promoter, a promoter that we show is substantially more active during growth on bacteria than the actin-6 promoter, which is widely used for axenic transformation. The procedure gives transformation frequencies of about 10(-5) with both strains Ax2 (capable of axenic growth) and NC4 (capable of growth only on bacteria). Using this vector, we have obtained NC4 strains carrying several beta-galactosidase reporter cassettes. Our vector can also be used in axenic transformations.
几乎所有用于转化社会变形虫盘基网柄菌的方法在至少部分过程中都依赖于无菌培养,即在合成培养基中生长。无菌培养需要多个突变。在这里,我们描述了一种可用于转化仅能在天然食物来源(细菌)上生长的野生型菌株的方法。该方法依赖于由V18启动子驱动的新选择盒,我们发现该启动子在细菌上生长时比广泛用于无菌转化的肌动蛋白-6启动子活性要强得多。该方法对Ax2(能够无菌生长)和NC4(仅能在细菌上生长)这两种菌株的转化频率约为10^(-5)。使用该载体,我们获得了携带多个β-半乳糖苷酶报告盒的NC4菌株。我们的载体也可用于无菌转化。