Jiao J, Douglas G R, Gingerich J D, Soper L M
Mutagenesis Section, Environmental Health Centre, Ottawa, Ontario, Canada.
Mutat Res. 1996 Nov 11;372(1):141-5. doi: 10.1016/S0027-5107(96)00181-9.
lacZ gene mutations in the transgenic Muta Mmouse can be detected by two different selection systems. While mutant frequencies recovered by phenyl-beta-D-galactoside (P-gal) selection are comparable with those obtained using 5-bromo-4-chloro-3-indolyl beta-D-galactopyranoside (X-gal) as substrate for beta-galactosidase, there may still be differences at the molecular level in the mutations detected by these two methods. Accordingly, we have examined a spectrum of mutants recovered from the X-gal system by regrowing these mutants on the P-gal plates. All colourless X-gal mutants grew normally on the P-gal plates. However, 11 out of 53 single light blue mutants, which express partial beta-galactosidase activity, produce few or no plaques on the P-gal plates, indicating the possible loss of some mutations using the positive selection system. Further analysis of mutant phenotypes and base changes indicates that such loss is not mutation-type specific. Our data suggest that the positive selection method can detect the majority of lacZ mutations detectable by visual selection and is more efficient at detecting mutants within a reduced range of beta-galactosidase activity.
转基因Muta M小鼠中的lacZ基因突变可通过两种不同的筛选系统进行检测。虽然通过苯基-β-D-半乳糖苷（P-gal）筛选获得的突变频率与使用5-溴-4-氯-3-吲哚基-β-D-吡喃半乳糖苷（X-gal）作为β-半乳糖苷酶底物所获得的频率相当，但这两种方法检测到的突变在分子水平上可能仍存在差异。因此，我们通过在P-gal平板上使这些突变体重生长，研究了从X-gal系统中回收的一系列突变体。所有无色的X-gal突变体在P-gal平板上均正常生长。然而，53个表达部分β-半乳糖苷酶活性的浅蓝色单突变体中有11个在P-gal平板上产生很少或不产生噬菌斑，这表明使用阳性筛选系统可能会丢失一些突变。对突变体表型和碱基变化的进一步分析表明，这种丢失并非突变类型特异性的。我们的数据表明，阳性筛选方法可以检测到通过视觉筛选可检测到的大多数lacZ突变，并且在检测β-半乳糖苷酶活性降低范围内的突变体时更有效。
