菌落原位杂交法筛选特定菌种的方法初探

Optimization of Colony in Situ Hybridization for Screening Specific Strains

菌落原位杂交法是重组DNA技术中最常用的方法之一,但该方法存在着杂交背景复杂及杂交结果不稳定性等缺点,所以进一步优化杂交体系显得尤为迫切。本研究通过优化杂交体系,最终确定利用溶菌酶处理杂交膜和采用非严紧型洗脱方式洗膜,不仅使得杂交体系的杂交结信号清晰,而且杂交背景较干净。研究结果表明,在探针特异性好的基础上,采用优化的菌落原位杂交体系筛选特定菌种是可行的。

Colony in situ Hybridization is one of the most widely-used methods in recombinant DNA technology. However, it is currently facing difficulties such as deep hybrid background and instability of results in hybridization. Therefore, the optimization of colony in situ hybridization system is necessary. In this research, through optimizing colony in situ hybridization system, we found that using lysozyme treatment system and the elution way of relaxation type for colony in situ hybridization, not only made positive dark dots distinguished clearly, but also resulted in clearer hybrid background. The results indicated that the use of colony in situ optimized hybridization system is feasible to screen specific strains if oligonucleotide probe is specific.