Harashima S, Takagi A, Oshima Y
Mol Cell Biol. 1984 Apr;4(4):771-8. doi: 10.1128/mcb.4.4.771-778.1984.
The frequency of cell fusion during transformation of yeast protoplasts with various yeast plasmids with a chromosome replicon (YRp or YCp) or 2 mu DNA (YEp) was estimated by two methods. In one method, a mixture of protoplasts of two haploid strains with identical mating type and complementary auxotrophic nuclear markers with or without cytoplasmic markers was transformed. When the number of various phenotypic classes of transformants for the nuclear markers was analyzed by equations derived from binominal distribution theory, the frequency of nuclear fusion among the transformants was 42 to 100% in transformations with the YRp or YCp plasmids and 28 to 39% with the YEp plasmids. In another method, a haploid bearing the sir mutation, which allows a diploid (or polyploid) homozygous for the MAT (mating type) locus to sporulate by the expression of the silent mating-type loci HML and HMR, was transformed with the plasmids. Sporulation ability was found in 43 to 95% of the transformants with the YRp or YCp plasmids, and 26 to 31% of the YEp transformants. When cytoplasmic mixing was included with the nuclear fusion, 96 to 100% of the transformants were found to be cell fusants. Based upon these observations, we concluded that transformation of yeast protoplasts is directly associated with cell fusion.
通过两种方法估算了用带有染色体复制子（YRp或YCp）或2μm DNA（YEp）的各种酵母质粒转化酵母原生质体过程中细胞融合的频率。在一种方法中，用具有相同交配型和互补营养缺陷型核标记（有或无细胞质标记）的两个单倍体菌株的原生质体混合物进行转化。当通过二项分布理论推导的方程式分析核标记转化体的各种表型类别的数量时，用YRp或YCp质粒转化时，转化体中核融合的频率为42%至100%，用YEp质粒转化时为28%至39%。在另一种方法中，用携带sir突变的单倍体进行转化，该突变通过沉默交配型位点HML和HMR的表达使MAT（交配型）位点纯合的二倍体（或多倍体）能够形成孢子。发现用YRp或YCp质粒转化的转化体中有43%至95%具有孢子形成能力，YEp转化体中有26%至31%具有孢子形成能力。当将细胞质混合与核融合包括在内时，发现96%至100%的转化体是细胞融合体。基于这些观察结果，我们得出结论，酵母原生质体的转化与细胞融合直接相关。
