Hotchkiss R D, Gabor M H
Proc Natl Acad Sci U S A. 1980 Jun;77(6):3553-7. doi: 10.1073/pnas.77.6.3553.
Efficiently regenerated single colonies from mixed multiply auxotrophic Bacillus subtilis protoplasts, fused with polyethylene glycol, reveal colonies carrying each of the parent types (biparentals) and recombinant colonies. The latter appear in high yields (up to 1% of certain recombinant classes); the yield of biparentals may be as large as 10%, in the range of the indicated physical fusion events. Many of the biparentals are diploids although, contrary to expectation, they are not complementing prototrophs, but show precisely the phenotype of one (either one) of the parent strains. Extensive pedigree analysis and subcloning of diploid lines show that they can propagate with varying stability on the appropriate parental selective medium to reproduce diploid progeny, parental segregants, and late-appearing recombinants, including some prototrophs. Thus, the principal product of intertype protoplast fusion is a diploid carrying two chromosomes, only one of which is expressed in each particular clone. The extinction of one parental genome is especially well demonstrated when it includes suppression of a normally dominant antibiotic sensitivity marker. In transformation experiments, DNA made from a selected diploid clone was able to transfer several of the unexpressed genes. The structural or topological character of DNA associated with the chromosome extinction remains unexplained.
从混合的多重营养缺陷型枯草芽孢杆菌原生质体中高效再生出单菌落，这些原生质体用聚乙二醇融合后，可揭示出携带每种亲本类型的菌落（双亲菌落）和重组菌落。后者产量很高（某些重组类型可达1%）；双亲菌落的产量可能高达10%，处于所示物理融合事件的范围内。许多双亲菌落是二倍体，尽管与预期相反，它们不是互补原养型，而是精确地表现出亲本菌株之一（任意一个）的表型。对二倍体系进行广泛的谱系分析和亚克隆表明，它们可以在合适的亲本选择培养基上以不同的稳定性繁殖，以产生二倍体后代、亲本分离株和后期出现的重组体，包括一些原养型。因此，不同类型原生质体融合的主要产物是携带两条染色体的二倍体，在每个特定克隆中只有其中一条染色体表达。当一个亲本基因组包括对正常显性抗生素敏感性标记的抑制时，该亲本基因组的消失尤其明显。在转化实验中，从选定的二倍体克隆制备的DNA能够转移几个未表达的基因。与染色体消失相关的DNA的结构或拓扑特征仍无法解释。
